Methods for the analysis of drugs. Methods for studying the quality of medicines Methods for analyzing drugs according to GF examples

The purpose of the study of medicinal substances is to establish the suitability of the medicinal product for medical use, i.e. compliance with its regulatory document for this drug.

Pharmaceutical analysis is the science of chemical characterization and measurement of biologically active substances at all stages of production: from the control of raw materials to the assessment of the quality of the resulting medicinal substance, the study of its stability, the establishment of expiration dates and the standardization of the finished dosage form. The peculiarities of pharmaceutical analysis are its versatility and variety of substances or their mixtures, including individual chemicals, complex mixtures of biological substances (proteins, carbohydrates, oligopeptides, etc.). Methods of analysis need to be constantly improved, and if chemical methods, including qualitative reactions, prevailed in the UP Pharmacopoeia, then at the present stage, mainly physicochemical and physical methods of analysis are used.

Pharmaceutical analysis, depending on the tasks, includes various aspects of drug quality control:
1. Pharmacopoeial analysis;
2. Stage-by-stage control of the production of medicines;
3. Analysis of individual drugs.

The main and most significant is the pharmacopoeial analysis, i.e. analysis of medicines for compliance with the standard - a pharmacopoeial monograph or other ND and, thus, confirmation of its suitability. Hence the requirements for high specificity, selectivity, accuracy and reliability of the analysis.

A conclusion about the quality of a medicinal product can only be made on the basis of a sample analysis (a statistically significant sample). The sampling procedure is indicated either in a private article or in a general article of the Global Fund X1 ed. (Issue 2) p.15. To test medicines for compliance with the requirements of regulatory and technical documentation, multi-stage sampling (sampling) is carried out. With multi-stage sampling, a sample (sample) is formed in stages and the products in each stage are selected randomly in proportional quantities from the units selected in the previous stage. The number of steps is determined by the type of packaging.

Stage 1: selection of packaging units (boxes, boxes, etc.);
Stage 2: selection of packaging units in packaging (boxes, bottles, cans, etc.);
Stage 3: selection of products in primary packaging (ampoules, vials, blisters, etc.).

To calculate the selection of the number of products at each stage, use the formula:

where n- the number of packaging units of this stage.

The specific sampling procedure is described in detail in the GF X1 edition, issue 2. In this case, the analysis is considered reliable if at least four samples are reproducible.

Pharmaceutical Analysis Criteria

For various purposes of the analysis, such criteria as the selectivity of the analysis, sensitivity, accuracy, the time of the analysis, the amount of the test substance are important.

The selectivity of the analysis is essential in the analysis of complex preparations consisting of several active components. In this case, the selectivity of the analysis is very important for the quantitative determination of each of the substances.

Requirements for accuracy and sensitivity depend on the object and purpose of the study. When testing for purity or impurities, highly sensitive methods are used. For stepwise production control, the time factor spent on analysis is important.

An important parameter of the analysis method is the sensitivity limit of the method. This limit means the lowest content at which a given substance can be reliably detected. The least sensitive are chemical methods of analysis and qualitative reactions. The most sensitive enzymatic and biological methods to detect single macromolecules of substances. Of those actually used, the most sensitive are radiochemical, catalytic and fluorescent methods, which make it possible to determine up to 10 -9%; sensitivity of spectrophotometric methods 10 -3 -10 -6%; potentiometric 10 -2%.

The term "analysis accuracy" simultaneously includes two concepts: reproducibility and correctness of the results obtained.

Reproducibility - characterizes the dispersion of the results of the analysis compared to the average value.

Correctness - reflects the difference between the actual and found content of the substance. The accuracy of the analysis depends on the quality of the instruments, the experience of the analyst, etc. The accuracy of the analysis cannot be higher than the accuracy of the least accurate measurement. This means that if the titration is accurate to ±0.2 ml plus leakage error is also ±0.2 ml, i.e. in total ±0.4 ml, then when 20 ml of titrant is consumed, the error is 0.2%. With a decrease in the sample and the amount of titrant, the accuracy decreases. Thus, titrimetric analysis allows determination with a relative error of ± (0.2-0.3)%. Each method has its own accuracy. When analyzing, it is important to have an understanding of the following concepts:

Gross mistakes- are a miscalculation of the observer or a violation of the analysis methodology. Such results are discarded as unreliable.

Systematic errors - reflect the correctness of the results of the analysis. They distort the measurement results, as a rule, in one direction by some constant value. Systematic errors can be partially eliminated by introducing corrections, instrument calibration, etc.

Random errors - reflect the reproducibility of the results of the analysis. They are called by uncontrolled variables. The arithmetic mean of random errors tends to zero. Therefore, for calculations, it is necessary to use not the results of single measurements, but the average of several parallel determinations.

Absolute error- represents the difference between the result obtained and the true value. This error is expressed in the same units as the value being determined.

Relative error definition is equal to the ratio of the absolute error to the true value of the determined value. It is usually expressed as a percentage or percentage.

The values ​​of relative errors depend on the method by which the analysis is performed and what the analyzed substance is - an individual substance and a mixture of many components.

The relative error in the study of individual substances by the spectrophotometric method is 2-3%, by IR spectrophotometry - 5-12%; liquid chromatography 3-4%; potentiometry 0.3-1%. Combined methods usually reduce the accuracy of the analysis. Biological methods are the least accurate - their relative error reaches 50%.

Methods for the identification of medicinal substances.

The most important indicator in the testing of medicinal substances is their identification or, as is customary in pharmacopoeial articles, authenticity. Numerous methods are used to determine the authenticity of medicinal substances. All the main and general are described in the GF X1 edition, issue 1. Historically, the main emphasis has been on chemical, incl. qualitative color reactions characterizing the presence of certain ions or functional groups in organic compounds, at the same time, physical methods were also widely used. In modern pharmacopoeias, the emphasis is on physico-chemical methods.

Let's focus on the main physical methods.

A fairly stable constant characterizing a substance, its purity and authenticity is the melting point. This indicator is widely used for the standardization of substances of medicinal substances. Methods for determining the melting point are described in detail in the GF X1, you yourself could try it out in laboratory classes. A pure substance has a constant melting point, however, when impurities are added to it, the melting point, as a rule, decreases very significantly. This effect is called a mixing test, and it is the mixing test that allows you to establish the authenticity of the drug in the presence of a standard sample or a known sample. There are, however, exceptions, as racemic sulphocamphoric acid melts at a higher temperature, and the various crystalline forms of indomethacin differ in melting point. Those. this method is one of the indicators that characterize both the purity of the product and its authenticity.

For some drugs, such an indicator as the solidification temperature is used. Another indicator characterizing a substance is the boiling point or temperature limits of distillation. This indicator characterizes liquid substances, for example, ethyl alcohol. The boiling point is a less characteristic indicator, it strongly depends on the pressure of the atmosphere, the possibility of the formation of mixtures or azeotropes and is used quite rarely.

Among other physical methods, it should be noted the determination density, viscosity. Standard methods of analysis are described in SP X1. The method that characterizes the authenticity of the drug is also the determination of its solubility in various solvents. According to GF X1 ed. This method is characterized as a property that can serve as an indicative characteristic of the test product. Along with the melting point, the solubility of a substance is one of the parameters by which the authenticity and purity of almost all medicinal substances are established. The pharmacopeia establishes an approximate gradation of substances by solubility from very easily soluble to practically insoluble. In this case, a substance is considered to be dissolved, in the solution of which no particles of the substance are observed in transmitted light.

Physical and chemical methods for determining authenticity.

The most informative in terms of determining the authenticity of substances are physicochemical methods based on the properties of the molecules of substances to interact with any physical factors. Physical and chemical methods include:

1.Spectral methods
UV spectroscopy
Spectroscopy in visible light
IR spectroscopy
Fluorescence spectroscopy
Atomic absorption spectroscopy
X-ray methods of analysis
Nuclear magnetic resonance
X-ray diffraction analysis

2. Sorption methods of analysis
Thin layer chromatography
Gas-liquid chromatography
High Performance Liquid Chromatography
Electrophoresis
Iontophoresis
Gel chromatography

3.Mass methods of analysis
Mass spectrometry
Chromatomass spectrometry

4. Electrochemical methods of analysis
Polarography
Electron paramagnetic resonance

5. Use of standard samples

Let us briefly consider the methods of analysis applicable in pharmacy. All these methods of analysis will be read to you in detail at the end of December by Professor V.I. Myagkikh. Some spectral methods are used to determine the authenticity of medicinal substances. The most reliable is the use of the low-frequency region of IR spectroscopy, where the absorption bands most reliably reflect this substance. I also call this area the fingerprint area. As a rule, comparison of IR spectra taken under standard conditions of a standard sample and a test sample is used to confirm authenticity. The coincidence of all absorption bands confirms the authenticity of the drug. The use of UV and visible spectroscopy is less reliable, because the nature of the spectrum is not individual and reflects only a certain chromophore in the structure of an organic compound. Atomic absorption spectroscopy and X-ray spectroscopy are used to analyze inorganic compounds, to identify chemical elements. Nuclear magnetic resonance makes it possible to establish the structure of organic compounds and is a reliable method for authenticating, however, due to the complexity of the instruments and the high cost, it is used very rarely and, as a rule, only for research purposes. Fluorescence spectroscopy is applicable only to a certain class of substances that fluoresce when exposed to UV radiation. In this case, the fluorescence spectrum and the fluorescence excitation spectrum are quite individual, but strongly depend on the medium in which the given substance is dissolved. This method is more commonly used for quantitation, especially of small quantities, as it is one of the most sensitive.

X-ray diffraction analysis is the most reliable method for confirming the structure of a substance, it allows you to establish the exact chemical structure of a substance, however, it is simply not suitable for stream analysis of authenticity and is used exclusively for scientific purposes.

Sorption methods of analysis found a very wide application in pharmaceutical analysis. They are used to determine authenticity, the presence of impurities, and quantification. You will be given a lecture in detail about these methods and the equipment used by Professor V.I. Myagkikh, a regional representative of Shimadzu, one of the main manufacturers of chromatographic equipment. These methods are based on the principle of sorption-desorption of substances on certain carriers in a carrier stream. Depending on the carrier and sorbent, they are divided into thin-layer chromatography, liquid column (analytical and preparative, including HPLC), gas-liquid chromatography, gel filtration, iontophoresis. The last two methods are used to analyze complex protein objects. A significant drawback of the methods is their relativity, i.e. Chromatography can characterize a substance and its quantity only when compared with a standard substance. However, it should be noted as a significant advantage - the high reliability of the method and accuracy, because. in chromatography, any mixture must be separated into individual substances and the result of the analysis is precisely the individual substance.

Mass spectrometric and electrochemical methods are rarely used to confirm authenticity.

A special place is occupied by methods for determining authenticity in comparison with a standard sample. This method is used quite widely in foreign pharmacopoeias to determine the authenticity of complex macromolecules, complex antibiotics, some vitamins, and other substances containing especially chiral carbon atoms, since it is difficult or even impossible to determine the authenticity of an optically active substance by other methods. A standard sample should be developed and issued on the basis of a developed and approved pharmacopoeial monograph. In Russia, only a few standard samples exist and are used, and the so-called RSOs are most often used for analysis - working standard samples prepared immediately before the experiment from known substances or corresponding substances.

Chemical methods of authentication.

The identification of medicinal substances by chemical methods is used mainly for inorganic medicinal substances, since other methods are most often not available or they require complex and expensive equipment. As already mentioned, inorganic elements are easily identified by atomic absorption or X-ray spectroscopy. Our Pharmacopoeia Monographs usually use chemical authentication methods. These methods are usually divided into the following:

Precipitation reactions of anions and cations. Typical examples are the precipitation reactions of sodium and potassium ions with (zincuranyl acetate and tartaric acid), respectively:

Such reactions are used in great variety and they will be discussed in detail in a special section of pharmaceutical chemistry regarding inorganic substances.

Redox reactions.

Redox reactions are used to reduce metals from oxides. For example, silver from its formalin oxide (silver mirror reaction):

The oxidation reaction of diphenylamine is the basis for testing the authenticity of nitrates and nitrites:

Reactions of neutralization and decomposition of anions.

Carbonates and hydrocarbonates under the action of mineral acids form carbonic acid, which decomposes to carbon dioxide:

Similarly, nitrites, thiosulfates, and ammonium salts decompose.

Changes in the color of a colorless flame. Sodium salts color the flame yellow, copper green, potassium purple, calcium brick red. It is this principle that is used in atomic absorption spectroscopy.

Decomposition of substances during pyrolysis. The method is used for preparations of iodine, arsenic, mercury. Of the currently used, the reaction of basic bismuth nitrate is most characteristic, which decomposes when heated to form nitrogen oxides:

Identification of organoelement medicinal substances.

Qualitative elemental analysis is used to identify compounds containing arsenic, sulfur, bismuth, mercury, phosphorus, and halogens in an organic molecule. Since the atoms of these elements are not ionized, preliminary mineralization is used to identify them, either by pyrolysis, or again by pyrolysis with sulfuric acid. Sulfur is determined by hydrogen sulfide reaction with potassium nitroprusside or lead salts. Iodine is also determined by pyrolysis by the release of elemental iodine. Of all these reactions, the identification of arsenic is of interest, not so much as a drug - they are practically not used, but as a method for monitoring impurities, but more on that later.

Testing the authenticity of organic medicinal substances. The chemical reactions used to test the authenticity of organic medicinal substances can be divided into three main groups:
1. General chemical reactions of organic compounds;
2. Reactions of formation of salts and complex compounds;
3. Reactions used to identify organic bases and their salts.

All these reactions are ultimately based on the principles of functional analysis, i.e. the reactive center of the molecule, which, when reacting, gives the appropriate response. Most often, this is a change in some properties of a substance: color, solubility, state of aggregation, etc.

Let us consider some examples of the use of chemical reactions for the identification of medicinal substances.

1. Reactions of nitration and nitrosation. They are used quite rarely, for example, to identify phenobarbital, phenacetin, dicain, although these drugs are almost never used in medical practice.

2. Diazotization and azo coupling reactions. These reactions are used to open primary amines. Diazotized amine combines with beta-naphthol to give a characteristic red or orange color.

3. Halogenation reactions. Used to open aliphatic double bonds - when bromine water is added, bromine is added to the double bond and the solution becomes colorless. A characteristic reaction of aniline and phenol is that when they are treated with bromine water, a tribromo derivative is formed, which precipitates.

4. Condensation reactions of carbonyl compounds. The reaction consists in the condensation of aldehydes and ketones with primary amines, hydroxylamine, hydrazines and semicarbazide:

The resulting azomethines (or Schiff bases) have a characteristic yellow color. The reaction is used to identify, for example, sulfonamides. The aldehyde used is 4-dimethylaminobenzaldehyde.

5. Oxidative condensation reactions. The process of oxidative cleavage and the formation of azomethine dye underlies ninhydrin reaction. This reaction is widely used for the discovery and photocolorimetric determination of α- and β-amino acids, in the presence of which an intense dark blue color appears. It is due to the formation of a substituted salt of diketohydrindylidene diketohydramine, a condensation product of excess ninhydrin and reduced ninhydrin with ammonia released during the oxidation of the test amino acid:

To open phenols, the reaction of the formation of triarylmethane dyes is used. So phenols interacting with formaldehyde form dyes. Similar reactions include the interaction of resorcinol with phthalic anhydride leading to the formation of a fluorescent dye - fluorescein.

Many other reactions are also used.

Of particular interest are reactions with the formation of salts and complexes. Inorganic salts of iron (III), copper (II), silver, cobalt, mercury (II) and others for testing the authenticity of organic compounds: carboxylic acids, including amino acids, derivatives of barbituric acid, phenols, sulfonamides, some alkaloids. The formation of salts and complex compounds occurs according to the general scheme:

R-COOH + MX = R-COOM + HX

The complex formation of amines proceeds similarly:

R-NH 2 + X = R-NH 2 X

One of the most common reagents in pharmaceutical analysis is a solution of iron (III) chloride. Interaction with phenols, it forms a colored solution of phenoxides, they are colored blue or purple. This reaction is used to discover phenol or resorcinol. However, meta-substituted phenols do not form colored compounds (thymol).

Copper salts form complex compounds with sulfonamides, cobalt salts with barbiturates. Many of these reactions are also used for quantitative determination.

Identification of organic bases and their salts. This group of methods is most often used in ready-made forms, especially in the study of solutions. So salts of organic amines, when alkalis are added, form a precipitate of a base (for example, a solution of papaverine hydrochloride) and vice versa, salts of organic acids, when a mineral acid is added, give a precipitate of an organic compound (for example, sodium salicylate). To identify organic bases and their salts, the so-called precipitation reagents are widely used. More than 200 precipitating reagents are known, which form water-insoluble simple or complex salts with organic compounds. The most commonly used solutions are given in the second volume of the SP 11th edition. An example is:
Scheibler's reagent - phosphotungstic acid;
Picric acid
Styphnic acid
Picramic acid

All these reagents are used for the precipitation of organic bases (for example, nitroxoline).

It should be noted that all these chemical reactions are used for the identification of medicinal substances not by themselves, but in combination with other methods, most often physicochemical, such as chromatography, spectroscopy. In general, it is necessary to pay attention to the fact that the problem of the authenticity of medicinal substances is a key one, because this fact determines the harmlessness, safety and effectiveness of the drug, so this indicator needs to be given great attention and it is not enough to confirm the authenticity of the substance by one method.

General requirements for purity tests.

Another equally important indicator of the quality of a medicinal product is purity. All medicinal products, regardless of the method of their preparation, are tested for purity. This determines the content of impurities in the preparation. It is conditionally possible to divide impurities into two groups: the first, impurities that have a pharmacological effect on the body; the second, impurities, indicating the degree of purification of the substance. The latter do not affect the quality of the drug, but in large quantities reduce its dose and, accordingly, reduce the activity of the drug. Therefore, all pharmacopoeias set certain limits for these impurities in drugs. Thus, the main criterion for the good quality of the drug is the absence of impurities, which is impossible by nature. The concept of the absence of impurities is associated with the detection limit of one method or another.

The physical and chemical properties of substances and their solutions give an approximate idea of ​​the presence of impurities in drugs and regulate their suitability for use. Therefore, in order to assess good quality, along with the establishment of authenticity and determination of the quantitative content, a number of physical and chemical tests are carried out to confirm the degree of its purity:

Transparency and degree of turbidity carried out by comparison with a turbidity standard, and transparency is determined by comparison with a solvent.

Chromaticity. A change in the degree of color may be due to:
a) the presence of an extraneous colored impurity;
b) a chemical change in the substance itself (oxidation, interaction with Me +3 and +2, or other chemical processes occurring with the formation of colored products. For example:

Resorcinol turns yellow during storage due to oxidation under the action of atmospheric oxygen to form quinones. In the presence of, for example, iron salts, salicylic acid acquires a purple color due to the formation of iron salicylates.

Color assessment is carried out by comparing the main experience with color standards, and colorlessness is determined by comparison with a solvent.

Very often, a test based on their interaction with concentrated sulfuric acid, which can act as an oxidizing or dehydrating agent, is used to detect organic impurities. As a result of such reactions, colored products are formed. The intensity of the resulting color should not exceed the corresponding color standard.

Determination of the degree of whiteness of powdered drugs– physical method, first included in GF X1. The degree of whiteness (hue) of solid medicinal substances can be assessed by various instrumental methods based on the spectral characteristics of the light reflected from the sample. To do this, reflectances are used when the sample is illuminated with white light obtained from a special source, with a spectral distribution or passed through light filters (with a transmission max of 614 nm (red) or 439 nm (blue)). You can also measure the reflectance of light passed through a green filter.

A more accurate assessment of the whiteness of medicinal substances can be carried out using reflection spectrophotometers. The value of the degree of whiteness and the degree of brightness are characteristics of the quality of whites and whites with shades of medicinal substances. Their permissible limits are regulated in private articles.

Determination of acidity, alkalinity, pH.

The change in these indicators is due to:
a) a change in the chemical structure of the medicinal substance itself:

b) the interaction of the drug with the container, for example, exceeding the permissible limits of alkalinity in a novocaine solution due to glass leaching;
c) absorption of gaseous products (CO 2 , NH 3) from the atmosphere.

Determination of the quality of medicines according to these indicators is carried out in several ways:

a) by changing the color of the indicator, for example, an admixture of mineral acids in boric acid is determined by methyl red, which does not change its color from the action of weak boric acid, but turns pink if it contains impurities of mineral acids.

b) titrimetric method - for example, to establish the permissible limit for the content of hydriodic acid formed during storage of a 10% alcohol solution of I 2, titration is carried out with alkali (no more than 0.3 ml of 0.1 mol / l NaOH by volume of the titrant). (Formaldehyde solution - titrated with alkali in the presence of phenolphthalein).

In some cases, the Global Fund sets the volume of titrant to determine the acidity or alkalinity.

Sometimes two titrated solutions are added in succession: first an acid and then an alkali.

c) by determining the pH value - for a number of drugs (and necessarily for all injection solutions) according to the NTD, it is envisaged to determine the pH value.

Techniques for preparing a substance in the study of acidity, alkalinity, pH

1. Preparation of a solution of a certain concentration specified in the NTD (for substances soluble in water)
2. For those insoluble in water, a suspension of a certain concentration is prepared and the acid-base properties of the filtrate are determined.
3. For liquid preparations immiscible with water, agitation with water is carried out, then the aqueous layer is separated and its acid-base properties are determined.
4. For insoluble solids and liquids, the determination can be carried out directly in suspension (ZnO)

The pH value approximately (up to 0.3 units) can be determined using indicator paper or a universal indicator.

The colorimetric method is based on the property of indicators to change their color at certain ranges of pH values. To perform the tests, buffer solutions with a constant concentration of hydrogen ions are used, differing from each other by a pH value of 0.2. To a series of such solutions and to the test solution add the same amount (2-3 drops) of the indicator. According to the coincidence of color with one of the buffer solutions, the pH value of the medium of the test solution is judged.

Determination of volatile substances and water.

Volatile substances can enter drugs either due to poor purification from solvents or intermediates, or as a result of the accumulation of degradation products. Water in the medicinal substance can be contained in the form of capillary, absorbed bound, chemically bound (hydrated and crystalline) or free.

Drying, distillation and titration with Fischer's solution are used to determine volatile substances and water.

drying method. The method is used to determine the loss in weight on drying. Losses can be due to the content of hygroscopic moisture and volatile substances in the substance. Dried in a bottle to constant weight at a certain temperature. More often, the substance is kept at a temperature of 100-105 ºС, but the conditions for drying and bringing to a constant mass may be different.

The determination of volatile substances can be carried out for some products by the method of ignition. The substance is heated in a crucible until the volatile substances are completely removed. then gradually increase the temperature until complete calcination at red heat. For example, the GPC regulates the determination of sodium carbonate impurities in the sodium bicarbonate medicinal substance by the calcination method. Sodium bicarbonate decomposes into sodium carbonate, carbon dioxide and water:

Theoretically, the weight loss is 36.9%. According to GPC, the loss in mass should be at least 36.6%. The difference between the theoretical and specified in the GPC mass loss determines the allowable limit of sodium carbonate impurities in the substance.

distillation method in GF 11 is called "Definition of water", it allows you to determine hygroscopic water. This method is based on the physical property of the vapors of two immiscible liquids. A mixture of water and an organic solvent distills at a lower temperature than either of these liquids. GPC1 recommends using toluene or xylene as the organic solvent. The water content in the test substance is determined by its volume in the receiver after the end of the distillation process.

Titration with Fisher's reagent. The method allows to determine the total content of both free and crystalline water in organic, inorganic substances, solvents. The advantage of this method is the speed of execution and selectivity with respect to water. Fisher's solution is a solution of sulfur dioxide, iodine and pyridine in methanol. Among the disadvantages of the method, in addition to the need for strict adherence to tightness, is the impossibility of determining water in the presence of substances that react with the components of the reagent.

Ash definition.

The ash content is due to mineral impurities that appear in organic substances in the process of obtaining auxiliary materials and equipment from the initial products (primarily metal cations), i.e. characterizes the presence of inorganic impurities in organic substances.

but) total ash- is determined by the results of combustion (ashing, mineralization) at high temperature, characterizes the sum of all inorganic substances-impurities.

Ash composition:
Carbonates: CaCO 3, Na 2 CO 3, K 2 CO 3, PbCO 3
Oxides: CaO, PbO
Sulphates: CaSO4
Chlorides: CaCl 2
Nitrates: NaNO 3

When obtaining medicines from plant materials, mineral impurities can be caused by dust pollution of plants, absorption of trace elements and inorganic compounds from soil, water, etc.

b) Ash insoluble in hydrochloric acid, obtained after treatment of total ash with dilute HCl. The chemical composition of the ash is heavy metal chlorides (AgCl, HgCl 2, Hg 2 Cl 2), i.e. highly toxic impurities.

in) sulfate ash- Sulphated ash is determined in assessing the good quality of many organic substances. Characterizes impurities Mn + n in a stable sulfate form. The resulting sulfate ash (Fe 3 (SO 4) 2, PbSO 4, CaSO 4) is used for the subsequent determination of heavy metal impurities.

Impurities of inorganic ions - C1 -, SO 4 -2, NH 4 +, Ca +2, Fe +3 (+2) , Pv +2, As +3 (+5)

Impurities:
a) impurities of a toxic nature (an admixture of CN - in iodine),
b) having an antagonistic effect (Na and K, Mg and Ca)

The absence of impurities that are not allowed in the medicinal substance is determined by a negative reaction with the appropriate reagents. Comparison in this case is carried out with a part of the solution, to which all reagents are added, except for the main one that opens this impurity (control experiment). A positive reaction indicates the presence of an impurity and the poor quality of the drug.

Permissible impurities - impurities that do not affect the pharmacological effect and the content of which is allowed in small quantities established by the NTD.

To establish the permissible limit for the content of ion impurities in medicines, reference solutions are used that contain the corresponding ion in a certain concentration.

Some medicinal substances are tested for the presence of impurities by titration, for example, the determination of the impurity of norsulfazole in the drug fthalazole. The admixture of norsulfazole in phthalazole is determined quantitatively by nitritometrically. Titration of 1 g of phthalazole should consume no more than 0.2 ml of 0.1 mol/l NaNO 2 .

General requirements for reactions that are used in tests for acceptable and unacceptable impurities:
1. sensitivity,
2. specificity,
3. reproducibility of the reaction used.

The results of reactions proceeding with the formation of colored products are observed in reflected light on a dull white background, and white precipitates in the form of turbidity and opalescence are observed in transmitted light on a black background.

Instrumental methods for determining impurities.

With the development of analysis methods, the requirements for the purity of medicinal substances and dosage forms are constantly increasing. In modern pharmacopoeias, along with the considered methods, various instrumental methods are used, based on the physicochemical, chemical and physical properties of substances. The use of UV and visible spectroscopy rarely gives positive results and this is due to the fact that the structure of impurities, especially organic drugs, as a rule. It is close to the structure of the drug itself, so the absorption spectra differ little, and the impurity concentration is usually tens of times lower than that of the main substance, which makes differential analysis methods unsuitable and allows one to estimate the impurity only approximately, i.e. as it is commonly called semi-quantitatively. The results are somewhat better if one of the substances, especially the impurity, forms a complex compound, while the other does not, then the maxima of the spectra differ significantly and it is already possible to determine the impurities quantitatively.

In recent years, IR-Fourier instruments have appeared at enterprises that allow determining both the content of the main substance and impurities, especially water, without destroying the sample, but their use is constrained by the high cost of instruments and the lack of standardized analysis methods.

Excellent impurity results are possible when the impurity fluoresces under UV light. The accuracy of such assays is very high, as is their sensitivity.

Wide application for testing for purity and quantitative determination of impurities both in medicinal substances (substances) and in dosage forms, which, perhaps, is no less important, because. many impurities are formed during the storage of drugs, obtained by chromatographic methods: HPLC, TLC, GLC.

These methods make it possible to determine impurities quantitatively, and each of the impurities individually, in contrast to other methods. The methods of HPLC and GLC chromatography will be discussed in detail in a lecture by prof. Myagkikh V.I. We will focus only on thin layer chromatography. The method of thin layer chromatography was discovered by the Russian scientist Tsvet and at the beginning existed as chromatography on paper. Thin layer chromatography (TLC) is based on the difference in the speeds of movement of the components of the analyzed mixture in a flat thin layer of the sorbent when the solvent (eluent) moves through it. Sorbents are silica gel, alumina, cellulose. Polyamide, eluents - organic solvents of different polarity or their mixtures with each other and sometimes with solutions of acids or alkalis and salts. The separation mechanism is due to the distribution coefficients between the sorbent and the liquid phase of the substance under study, which in turn is associated with many, including the chemical and physicochemical properties of the substances.

In TLC, the surface of an aluminum or glass plate is covered with a sorbent suspension, dried in air, and activated to remove traces of solvent (moisture). In practice, commercially manufactured plates with a fixed layer of sorbent are usually used. Drops of the analyzed solution with a volume of 1-10 μl are applied to the sorbent layer. The edge of the plate is immersed in the solvent. The experiment is carried out in a special chamber - a glass vessel, closed with a lid. The solvent moves through the layer under the action of capillary forces. Simultaneous separation of several different mixtures is possible. To increase the separation efficiency, multiple elution is used either in the perpendicular direction with the same or a different eluent.

After the completion of the process, the plate is dried in air and the position of the chromatographic zones of the components is set in various ways, for example, by irradiation with UV radiation, by spraying with coloring reagents, and kept in iodine vapor. On the resulting distribution pattern (chromatogram), the chromatographic zones of the mixture components are arranged in the form of spots in accordance with their sorbability in the given system.

The position of the chromatographic zones on the chromatogram is characterized by the value of R f . which is equal to the ratio of the path l i traversed by the i-th component from the starting point to the path Vп R f = l i / l.

The value of R f depends on the coefficient of distribution (adsorption) K і and the ratio of the volumes of the mobile (V p) and stationary (V n) phases.

Separation in TLC is affected by a number of factors: the composition and properties of the eluent, the nature, fineness and porosity of the sorbent, temperature, humidity, the size and thickness of the sorbent layer, and the dimensions of the chamber. Standardization of experimental conditions allows setting R f with a relative standard deviation of 0.03.

Identification of the components of the mixture is carried out by the values ​​of R f . The quantitative determination of substances in the zones can be carried out directly on the sorbent layer by the area of ​​the chromatographic zone, the fluorescence intensity of the component or its combination with a suitable reagent, by radiochemical methods. Automatic scanning instruments are also used to measure the absorption, transmission, reflection of light, or radioactivity of chromatographic zones. The separated zones can be removed from the plate together with the sorbent layer, the component can be desorbed into the solvent, and the solution can be analyzed spectrophotometrically. Using TLC, substances can be determined in quantities from 10 -9 to 10 -6; the error of determination is not less than 5-10%.

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• Introduction
• Chapter 1. Basic Principles of Pharmaceutical Analysis
• 1.1 Pharmaceutical analysis criteria
• 1.2 Errors in Pharmaceutical Analysis
• 1.4 Sources and causes of poor quality of medicinal substances
• 1.5 General requirements for purity tests
• 1.6 Methods of pharmaceutical analysis and their classification
• Chapter 2. Physical Methods of Analysis
• 2.1 Verification of physical properties or measurement of physical constants of drug substances
• 2.2 Setting the pH of the medium
• 2.3 Determination of clarity and turbidity of solutions
• 2.4 Estimation of chemical constants
• Chapter 3. Chemical Methods of Analysis
• 3.1 Features of chemical methods of analysis
• 3.2 Gravimetric (weight) method
• 3.3 Titrimetric (volumetric) methods
• 3.4 Gasometric analysis
• 3.5 Quantitative elemental analysis
• Chapter 4. Physical and chemical methods of analysis
• 4.1 Features of physicochemical methods of analysis
• 4.2 Optical methods
• 4.3 Absorption methods
• 4.4 Methods based on emission of radiation
• 4.5 Methods based on the use of a magnetic field
• 4.6 Electrochemical methods
• 4.7 Separation methods
• 4.8 Thermal methods of analysis
• Chapter 5
• 5.1 Biological quality control of medicines
• 5.2 Microbiological control of medicinal products
• conclusions
• List of used literature

Introduction

Pharmaceutical analysis is the science of chemical characterization and measurement of biologically active substances at all stages of production: from the control of raw materials to the assessment of the quality of the resulting medicinal substance, the study of its stability, the establishment of expiration dates and the standardization of the finished dosage form. Pharmaceutical analysis has its own specific features that distinguish it from other types of analysis. These features lie in the fact that substances of various chemical nature are subjected to analysis: inorganic, organoelement, radioactive, organic compounds from simple aliphatic to complex natural biologically active substances. The range of concentrations of analytes is extremely wide. The objects of pharmaceutical analysis are not only individual medicinal substances, but also mixtures containing a different number of components. The number of medicines is increasing every year. This necessitates the development of new methods of analysis.

Methods of pharmaceutical analysis need to be systematically improved due to the continuous increase in the requirements for the quality of drugs, and the requirements for both the degree of purity of medicinal substances and the quantitative content are growing. Therefore, it is necessary to widely use not only chemical, but also more sensitive physical and chemical methods for assessing the quality of drugs.

The requirements for pharmaceutical analysis are high. It should be sufficiently specific and sensitive, accurate in relation to the standards stipulated by GF XI, VFS, FS and other scientific and technical documentation, carried out in short periods of time using minimal quantities of tested drugs and reagents.

Pharmaceutical analysis, depending on the tasks, includes various forms of drug quality control: pharmacopoeial analysis, step-by-step control of the production of medicines, analysis of individual dosage forms, express analysis in a pharmacy, and biopharmaceutical analysis.

Pharmacopoeial analysis is an integral part of pharmaceutical analysis. It is a set of methods for studying drugs and dosage forms set forth in the State Pharmacopoeia or other regulatory and technical documentation (VFS, FS). Based on the results obtained during the pharmacopoeial analysis, a conclusion is made on the compliance of the medicinal product with the requirements of the Global Fund or other regulatory and technical documentation. In case of deviation from these requirements, the drug is not allowed to be used.

The conclusion about the quality of the medicinal product can only be made on the basis of the analysis of the sample (sample). The procedure for its selection is indicated either in a private article or in a general article of the Global Fund XI (issue 2). Sampling is carried out only from undamaged sealed and packed in accordance with the requirements of the NTD packaging units. At the same time, the requirements for precautionary measures for working with poisonous and narcotic drugs, as well as for toxicity, flammability, explosiveness, hygroscopicity and other properties of drugs, must be strictly observed. To test for compliance with the requirements of the NTD, multi-stage sampling is carried out. The number of steps is determined by the type of packaging. At the last stage (after control by appearance), a sample is taken in the amount necessary for four complete physical and chemical analyzes (if the sample is taken for controlling organizations, then for six such analyzes).

From the "angro" packaging, point samples are taken, taken in equal quantities from the top, middle and bottom layers of each packaging unit. After establishing homogeneity, all these samples are mixed. Loose and viscous drugs are taken with a sampler made of an inert material. Liquid medicinal products are thoroughly mixed before sampling. If this is difficult to do, then point samples are taken from different layers. The selection of samples of finished medicinal products is carried out in accordance with the requirements of private articles or control instructions approved by the Ministry of Health of the Russian Federation.

Performing a pharmacopoeial analysis allows you to establish the authenticity of the drug, its purity, to determine the quantitative content of the pharmacologically active substance or ingredients that make up the dosage form. While each of these stages has a specific purpose, they cannot be viewed in isolation. They are interrelated and complement each other. For example, melting point, solubility, pH of an aqueous solution, etc. are criteria for both authenticity and purity of a medicinal substance.

Chapter 1. Basic Principles of Pharmaceutical Analysis

1.1 Pharmaceutical analysis criteria

At various stages of pharmaceutical analysis, depending on the tasks set, such criteria as selectivity, sensitivity, accuracy, time spent on the analysis, and the amount of the analyzed drug (dosage form) are important.

The selectivity of the method is very important when analyzing mixtures of substances, since it makes it possible to obtain the true values ​​of each of the components. Only selective methods of analysis make it possible to determine the content of the main component in the presence of decomposition products and other impurities.

Requirements for the accuracy and sensitivity of pharmaceutical analysis depend on the object and purpose of the study. When testing the degree of purity of the drug, methods are used that are highly sensitive, allowing you to set the minimum content of impurities.

When performing step-by-step production control, as well as when conducting express analysis in a pharmacy, an important role is played by the time factor spent on the analysis. For this, methods are chosen that allow the analysis to be carried out in the shortest time intervals and at the same time with sufficient accuracy.

In the quantitative determination of a medicinal substance, a method is used that is distinguished by selectivity and high accuracy. The sensitivity of the method is neglected, given the possibility of performing an analysis with a large sample of the drug.

A measure of the sensitivity of a reaction is the limit of detection. It means the lowest content at which the presence of the determined component with a given confidence probability can be detected by this method. The term "limit of detection" was introduced instead of such a concept as "discovered minimum", it is also used instead of the term "sensitivity". The sensitivity of qualitative reactions is influenced by such factors as the volumes of solutions of reacting components, concentrations of reagents, pH of the medium, temperature, duration experience.This should be taken into account when developing methods for qualitative pharmaceutical analysis.To establish the sensitivity of reactions, the absorbance index (specific or molar) established by the spectrophotometric method is increasingly used.In chemical analysis, the sensitivity is set by the value of the limit of detection of a given reaction.Physicochemical methods are distinguished by high sensitivity The most highly sensitive are radiochemical and mass spectral methods, which make it possible to determine 10 -8 -10 -9% of the analyte, polarographic and fluorimetric 10 -6 -10 -9%, sensitivity of spectrophotometric methods is 10 -3 -10 -6 %, potentiometric 10 -2%.

The term "analysis accuracy" simultaneously includes two concepts: reproducibility and correctness of the obtained results. Reproducibility characterizes the scatter of the results of an analysis compared to the mean. Correctness reflects the difference between the actual and found content of the substance. The accuracy of the analysis for each method is different and depends on many factors: the calibration of measuring instruments, the accuracy of weighing or measuring, the experience of the analyst, etc. The accuracy of the analysis result cannot be higher than the accuracy of the least accurate measurement.

So, when calculating the results of titrimetric determinations, the least accurate figure is the number of milliliters of titrant used for titration. In modern burettes, depending on their accuracy class, the maximum measurement error is about ±0.02 ml. The leakage error is also ±0.02 ml. If, with the indicated total measurement and leakage error of ±0.04 ml, 20 ml of titrant is consumed for titration, then the relative error will be 0.2%. With a decrease in the sample and the number of milliliters of titrant, the accuracy decreases accordingly. Thus, titrimetric determination can be performed with a relative error of ±(0.2--0.3)%.

The accuracy of titrimetric determinations can be improved by using microburettes, the use of which significantly reduces errors from inaccurate measurement, leakage, and temperature effects. An error is also allowed when taking a sample.

The weighing of the sample when performing the analysis of the medicinal substance is carried out with an accuracy of ± 0.2 mg. When taking a sample of 0.5 g of the drug, which is usual for pharmacopoeial analysis, and weighing accuracy of ± 0.2 mg, the relative error will be 0.4%. When analyzing dosage forms, performing express analysis, such accuracy when weighing is not required, therefore, a sample is taken with an accuracy of ± (0.001--0.01) g, i.e. with a limiting relative error of 0.1--1%. This can also be attributed to the accuracy of weighing the sample for colorimetric analysis, the accuracy of the results of which is ±5%.

1.2 Mistakes during Pharmaceutical Analysis

When performing a quantitative determination by any chemical or physico-chemical method, three groups of errors can be made: gross (misses), systematic (certain) and random (uncertain).

Gross errors are the result of a miscalculation of the observer when performing any of the determination operations or incorrectly performed calculations. Results with gross errors are discarded as poor quality.

Systematic errors reflect the correctness of the results of the analysis. They distort the measurement results, usually in one direction (positive or negative) by some constant value. The reason for systematic errors in the analysis may be, for example, the hygroscopicity of the drug when weighing its sample; imperfection of measuring and physico-chemical instruments; experience of the analyst, etc. Systematic errors can be partially eliminated by making corrections, instrument calibration, etc. However, it is always necessary to ensure that the systematic error is commensurate with the error of the instrument and does not exceed the random error.

Random errors reflect the reproducibility of the results of the analysis. They are called by uncontrolled variables. The arithmetic mean of random errors tends to zero when a large number of experiments are performed under the same conditions. Therefore, for calculations, it is necessary to use not the results of single measurements, but the average of several parallel determinations.

The correctness of the results of the determinations is expressed by the absolute error and the relative error.

The absolute error is the difference between the result obtained and the true value. This error is expressed in the same units as the determined value (grams, milliliters, percent).

The relative error of the determination is equal to the ratio of the absolute error to the true value of the quantity being determined. The relative error is usually expressed as a percentage (by multiplying the resulting value by 100). Relative errors in determinations by physicochemical methods include both the accuracy of performing preparatory operations (weighing, measuring, dissolving) and the accuracy of performing measurements on the device (instrumental error).

The values ​​of relative errors depend on the method used to perform the analysis and whether the analyzed object is an individual substance or a multicomponent mixture. Individual substances can be determined by analyzing the spectrophotometric method in the UV and visible regions with a relative error of ±(2--3)%, IR spectrophotometry ±(5--12)%, gas-liquid chromatography ±(3--3 ,five)%; polarography ±(2--3)%; potentiometry ±(0.3--1)%.

When analyzing multicomponent mixtures, the relative error of determination by these methods increases by about a factor of two. The combination of chromatography with other methods, in particular the use of chromato-optical and chromatoelectrochemical methods, makes it possible to analyze multicomponent mixtures with a relative error of ±(3--7)%.

The accuracy of biological methods is much lower than that of chemical and physicochemical methods. The relative error of biological determinations reaches 20-30 and even 50%. To improve accuracy, SP XI introduced a statistical analysis of the results of biological tests.

The relative determination error can be reduced by increasing the number of parallel measurements. However, these possibilities have a certain limit. It is advisable to reduce the random measurement error by increasing the number of experiments until it becomes less than the systematic error. Typically, 3-6 parallel measurements are performed in pharmaceutical analysis. When statistically processing the results of determinations, in order to obtain reliable results, at least seven parallel measurements are performed.

1.3 General principles for testing the identity of medicinal substances

Authenticity testing is a confirmation of the identity of the analyzed medicinal substance (dosage form), carried out on the basis of the requirements of the Pharmacopoeia or other regulatory and technical documentation (NTD). Tests are performed by physical, chemical and physico-chemical methods. An indispensable condition for an objective test of the authenticity of a medicinal substance is the identification of those ions and functional groups included in the structure of molecules that determine pharmacological activity. With the help of physical and chemical constants (specific rotation, pH of the medium, refractive index, UV and IR spectrum), other properties of molecules that affect the pharmacological effect are also confirmed. Chemical reactions used in pharmaceutical analysis are accompanied by the formation of colored compounds, the release of gaseous or water-insoluble compounds. The latter can be identified by their melting point.

1.4 Sources and causes of poor quality of medicinal substances

The main sources of technological and specific impurities are equipment, raw materials, solvents and other substances that are used in the preparation of medicines. The material from which the equipment is made (metal, glass) can serve as a source of impurities of heavy metals and arsenic. With poor cleaning, the preparations may contain impurities of solvents, fibers of fabrics or filter paper, sand, asbestos, etc., as well as acid or alkali residues.

The quality of synthesized medicinal substances can be influenced by various factors.

Technological factors are the first group of factors that influence the process of drug synthesis. The degree of purity of the starting materials, temperature, pressure, pH of the medium, solvents used in the synthesis process and for purification, mode and temperature of drying, fluctuating even within small limits - all these factors can lead to the appearance of impurities that accumulate from one to another stage. In this case, the formation of products of side reactions or decomposition products, the processes of interaction of the initial and intermediate synthesis products with the formation of such substances, from which it is difficult then to separate the final product, can occur. In the process of synthesis, the formation of various tautomeric forms is also possible both in solutions and in the crystalline state. For example, many organic compounds can exist in amide, imide, and other tautomeric forms. And quite often, depending on the conditions of preparation, purification and storage, the medicinal substance can be a mixture of two tautomers or other isomers, including optical ones, differing in pharmacological activity.

The second group of factors is the formation of various crystalline modifications, or polymorphism. About 65% of medicinal substances belonging to the number of barbiturates, steroids, antibiotics, alkaloids, etc., form 1-5 or more different modifications. The rest give during crystallization stable polymorphic and pseudopolymorphic modifications. They differ not only in physicochemical properties (melting point, density, solubility) and pharmacological action, but they have different values ​​of free surface energy and, consequently, unequal resistance to the action of air oxygen, light, moisture. This is caused by changes in the energy levels of molecules, which affects the spectral, thermal properties, solubility and absorption of drugs. The formation of polymorphic modifications depends on the crystallization conditions, the solvent used, and the temperature. The transformation of one polymorphic form into another occurs during storage, drying, grinding.

In medicinal substances obtained from plant and animal raw materials, the main impurities are associated natural compounds (alkaloids, enzymes, proteins, hormones, etc.). Many of them are very similar in chemical structure and physicochemical properties to the main extraction product. Therefore, cleaning it is very difficult.

The dustiness of industrial premises of chemical-pharmaceutical enterprises can have a great influence on the contamination with impurities of some drugs by others. In the working area of ​​these premises, provided that one or more preparations (dosage forms) are received, all of them can be contained in the form of aerosols in the air. In this case, the so-called "cross-contamination" occurs.

The World Health Organization (WHO) in 1976 developed special rules for the organization of production and quality control of medicines, which provide for the conditions for preventing "cross-contamination".

Not only the technological process, but also storage conditions are important for the quality of drugs. The good quality of preparations is affected by excessive moisture, which can lead to hydrolysis. As a result of hydrolysis, basic salts, saponification products and other substances with a different pharmacological action are formed. When storing crystalline preparations (sodium arsenate, copper sulfate, etc.), on the contrary, it is necessary to observe conditions that exclude the loss of crystallization water.

When storing and transporting drugs, it is necessary to take into account the effect of light and oxygen in the air. Under the influence of these factors, decomposition of, for example, substances such as bleach, silver nitrate, iodides, bromides, etc. can occur. Of great importance is the quality of the container used to store medicines, as well as the material from which it is made. The latter can also be a source of impurities.

Thus, impurities contained in medicinal substances can be divided into two groups: technological impurities, i.e. introduced by the feedstock or formed during the production process, and impurities acquired during storage or transportation, under the influence of various factors (heat, light, atmospheric oxygen, etc.).

The content of these and other impurities must be strictly controlled in order to exclude the presence of toxic compounds or the presence of indifferent substances in medicinal products in such quantities that interfere with their use for specific purposes. In other words, the medicinal substance must have a sufficient degree of purity, and therefore, meet the requirements of a certain specification.

A drug substance is pure if further purification does not change its pharmacological activity, chemical stability, physical properties and bioavailability.

In recent years, due to the deterioration of the environmental situation, medicinal plant raw materials are also tested for the presence of impurities of heavy metals. The importance of such tests is due to the fact that when conducting studies of 60 different samples of plant materials, the content of 14 metals was established in them, including such toxic ones as lead, cadmium, nickel, tin, antimony and even thallium. Their content in most cases significantly exceeds the established maximum allowable concentrations for vegetables and fruits.

The pharmacopoeial test for the determination of heavy metal impurities is one of the widely used in all national pharmacopoeias of the world, which recommend it for the study of not only individual medicinal substances, but also oils, extracts, and a number of injectable dosage forms. In the opinion of the WHO Expert Committee, such tests should be carried out on medicinal products having single doses of at least 0.5 g.

1.5 General requirements for purity tests

Evaluation of the degree of purity of a medicinal product is one of the important steps in pharmaceutical analysis. All drugs, regardless of the method of preparation, are tested for purity. At the same time, the content of impurities is determined. They can be divided into two groups: impurities that affect the pharmacological action of the drug, and impurities that indicate the degree of purification of the substance. The latter do not affect the pharmacological effect, but their presence in large quantities reduces the concentration and, accordingly, reduces the activity of the drug. Therefore, pharmacopoeias set certain limits for these impurities in drugs.

Thus, the main criterion for the good quality of a medicinal product is the presence of acceptable limits for physiologically inactive impurities and the absence of toxic impurities. The concept of absence is conditional and is associated with the sensitivity of the test method.

The general requirements for purity tests are the sensitivity, specificity and reproducibility of the reaction used, as well as the suitability of its use for establishing acceptable limits for impurities.

For purity tests, select reactions with a sensitivity that allows you to determine the acceptable limits of impurities in a given medicinal product. These limits are established by preliminary biological testing, taking into account the possible toxic effects of the impurity.

There are two ways to determine the maximum content of impurities in the test preparation (reference and non-reference). One of them is based on comparison with a reference solution (standard). At the same time, under the same conditions, a color or turbidity is observed that occurs under the action of any reagent. The second way is to set a limit on the content of impurities based on the absence of a positive reaction. In this case, chemical reactions are used, the sensitivity of which is lower than the detection limit of admissible impurities.

To speed up the performance of tests for purity, their unification and achieving the same accuracy of analysis in domestic pharmacopoeias, a system of standards was used. A reference is a sample containing a certain amount of an impurity to be discovered. The determination of the presence of impurities is carried out by the colorimetric or nephelometric method, comparing the results of reactions in the standard solution and in the drug solution after adding the same amounts of the corresponding reagents. The accuracy achieved in this case is quite sufficient to establish whether more or less impurities are contained in the test preparation than is permissible.

When performing tests for purity, it is necessary to strictly follow the general guidelines provided for by pharmacopoeias. Water and reagents used should not contain ions, the presence of which is established; test tubes should be of the same diameter and colorless; samples must be weighed to the nearest 0.001 g; reagents should be added simultaneously and in equal amounts to both the reference and the test solution; the resulting opalescence is observed in transmitted light against a dark background, and the color is observed in reflected light against a white background. If the absence of an impurity is established, then all reagents are added to the test solution, except for the main one; then the resulting solution is divided into two equal parts and the main reagent is added to one of them. When compared, there should be no noticeable differences between both parts of the solution.

It should be borne in mind that the sequence and rate of addition of the reagent will affect the results of the purity tests. Sometimes it is also necessary to observe the time interval during which the result of the reaction should be monitored.

The source of impurities in the production of finished dosage forms can be poorly purified fillers, solvents and other excipients. Therefore, the degree of purity of these substances must be carefully controlled before they are used in production.

1.6 Methods of pharmaceutical analysis and their classification

Pharmaceutical analysis uses a variety of research methods: physical, physico-chemical, chemical, biological. The use of physical and physico-chemical methods requires appropriate instruments and instruments, therefore, these methods are also called instrumental, or instrumental.

The use of physical methods is based on the measurement of physical constants, for example, transparency or degree of turbidity, color, humidity, melting, solidification and boiling points, etc.

With the help of physicochemical methods, the physical constants of the analyzed system are measured, which change as a result of chemical reactions. This group of methods includes optical, electrochemical, chromatographic.

Chemical methods of analysis are based on the performance of chemical reactions.

Biological control of medicinal substances is carried out on animals, individual isolated organs, groups of cells, on certain strains of microorganisms. Establish the strength of the pharmacological effect or toxicity.

Methods used in pharmaceutical analysis should be sensitive, specific, selective, fast and suitable for rapid analysis in a pharmacy setting.

Chapter 2. Physical Methods of Analysis

2.1 Verification of physical properties or measurement of physical constants of medicinal substances

The authenticity of the medicinal substance is confirmed; state of aggregation (solid, liquid, gas); color, smell; the shape of the crystals or the type of amorphous substance; hygroscopicity or degree of weathering in air; resistance to light, air oxygen; volatility, mobility, flammability (of liquids). The color of a medicinal substance is one of the characteristic properties that allows its preliminary identification.

Determination of the degree of whiteness of powdered medicines is a physical method, first included in the Global Fund XI. The degree of whiteness (hue) of solid medicinal substances can be estimated by various instrumental methods based on the spectral characteristics of the light reflected from the sample. To do this, measure the reflection coefficients when the sample is illuminated with white light obtained from a special source with a spectral distribution or passed through light filters with a maximum transmission of 614 nm (red) or 459 nm (blue). You can also measure the reflectance of light passed through a green filter (522 nm). The reflection coefficient is the ratio of the magnitude of the reflected light flux to the magnitude of the incident light flux. It allows you to determine the presence or absence of a color shade in medicinal substances by the degree of whiteness and degree of brightness. For white or white substances with a grayish tint, the degree of whiteness is theoretically equal to 1. Substances in which it is 0.95--1.00, and the degree of brightness< 0,85, имеют сероватый оттенок.

A more accurate assessment of the whiteness of medicinal substances can be carried out using reflection spectrophotometers, for example, SF-18, manufactured by LOMO (Leningrad Optical and Mechanical Association). The intensity of color or grayish shades is set according to the absolute reflection coefficients. Whiteness and brightness values are characteristics of the quality of whites and whites with hints of medicinal substances. Their permissible limits are regulated in private articles.

More objective is the establishment of various physical constants: melting (decomposition) temperature, solidification or boiling point, density, viscosity. An important indicator of authenticity is the solubility of the drug in water, solutions of acids, alkalis, organic solvents (ether, chloroform, acetone, benzene, ethyl and methyl alcohol, oils, etc.).

The constant characterizing the homogeneity of solids is the melting point. It is used in pharmaceutical analysis to establish the identity and purity of most drug solids. It is known that this is the temperature at which the solid is in equilibrium with the liquid phase when the vapor phase is saturated. The melting point is a constant value for an individual substance. The presence of even a small amount of impurities changes (as a rule, reduces) the melting point of a substance, which makes it possible to judge the degree of its purity. The identity of the compound under study can be confirmed by a mixed melting test, since a mixture of two substances having the same melting points melts at the same temperature.

To establish the melting point, SP XI recommends a capillary method that allows you to confirm the authenticity and approximately the degree of purity of the medicinal product. Since a certain content of impurities is allowed in medicinal preparations (normalized by FS or VFS), the melting point may not always be clearly expressed. Therefore, most pharmacopoeias, including SP XI, under the melting point mean the temperature range at which the process of melting of the test drug occurs from the appearance of the first drops of liquid to the complete transition of the substance into a liquid state. Some organic compounds decompose when heated. This process occurs at the decomposition temperature and depends on a number of factors, in particular on the heating rate.

The intervals of melting temperatures given in private articles of the State Pharmacopoeia (FS, VFS) indicate that the interval between the beginning and end of the melting of the medicinal substance should not exceed 2°C. If it exceeds 2°C, then the private article should indicate by what amount. If the transition of a substance from a solid to a liquid state is fuzzy, then instead of the melting temperature interval, the temperature is set at which only the beginning or only the end of melting occurs. This temperature value should fit into the interval given in the private article of the Global Fund (FS, VFS).

Description of the device and methods for determining the melting point is given in the SP XI, issue 1 (p. 16). Depending on the physical properties, various methods are used. One of them is recommended for solids that are easily powdered, and the other two are for substances that do not grind into powder (fats, wax, paraffin, petroleum jelly, etc.). It should be borne in mind that the accuracy of establishing the temperature interval at which the melting of the test substance occurs can be affected by the conditions of sample preparation, the rate of rise and accuracy of temperature measurement, and the experience of the analyst.

In GF XI, no. 1 (p. 18), the conditions for determining the melting point are specified and a new device with a measurement range of 20 to 360°C (PTP) with electric heating is recommended. It is distinguished by the presence of a glass heater block, which is heated by coiled constantan wire, an optical device and a control panel with a nomogram. The capillaries for this device should be 20 cm long. The PTP device provides a higher accuracy in determining the melting point. If discrepancies are obtained in determining the melting point (specified in a private article), then the results of its determination on each of the devices used should be given.

The solidification point is understood as the highest, remaining for a short time, constant temperature at which the transition of a substance from a liquid to a solid state occurs. In GF XI, no. 1 (p. 20) describes the design of the device and the method for determining the solidification temperature. Compared to GF X, an addition has been made to it regarding substances capable of supercooling.

The boiling point, or, more precisely, the temperature limits of distillation, is the interval between the initial and final boiling points at normal pressure of 760 mmHg. (101.3 kPa). The temperature at which the first 5 drops of liquid were distilled into the receiver is called the initial boiling point, and the temperature at which 95% of the liquid passed into the receiver is called the final boiling point. The indicated temperature limits can be set by the macromethod and the micromethod. In addition to the device recommended by GF XI, vol. 1 (p. 18), to determine the melting point (MTP), a device for determining the temperature limits of distillation (TPP) of liquids, manufactured by the Klin plant "Laborpribor" (SP XI, issue 1, p. 23), can be used. This instrument provides more accurate and reproducible results.

Keep in mind that the boiling point depends on atmospheric pressure. The boiling point is set only for a relatively small number of liquid drugs: cyclopropane, chloroethyl, ether, halothane, chloroform, trichlorethylene, ethanol.

When determining the density, the mass of a substance of a certain volume is taken. The density is set using a pycnometer or hydrometer according to the methods described in SP XI, vol. 1 (p. 24--26), strictly observing the temperature regime, since the density depends on temperature. This is usually achieved by thermostating the pycnometer at 20°C. Certain intervals of density values ​​confirm the authenticity of ethyl alcohol, glycerin, vaseline oil, vaseline, solid paraffin, halogen derivatives of hydrocarbons (chloroethyl, halothane, chloroform), formaldehyde solution, ether for anesthesia, amyl nitrite, etc. GF XI, issue. 1 (p. 26) recommends establishing the alcohol content in preparations of ethyl alcohol 95, 90, 70 and 40% by density, and in dosage forms either by distillation with subsequent determination of density, or by the boiling point of water-alcohol solutions (including tinctures).

Distillation is carried out by boiling certain amounts of alcohol-water mixtures (tinctures) in flasks hermetically connected to the receiver. The latter is a volumetric flask with a capacity of 50 ml. Collect 48 ml of distillate, bring its temperature to 20°C and add water to the mark. The distillation density is set with a pycnometer.

When determining alcohol (in tinctures) by boiling point, use the device described in SP XI, vol. 1 (p. 27). The thermometer readings are taken 5 minutes after the start of boiling, when the boiling point stabilizes (deviations are not more than ±0.1°C). The result obtained is converted to normal atmospheric pressure. The alcohol concentration is calculated using the tables available in GF XI, vol. 1 (p. 28).

Viscosity (internal friction) is a physical constant that confirms the authenticity of liquid medicinal substances. There are dynamic (absolute), kinematic, relative, specific, reduced and characteristic viscosity. Each of them has its own units of measurement.

To assess the quality of liquid preparations having a viscous consistency, for example, glycerin, petrolatum, oils, the relative viscosity is usually determined. It is the ratio of the viscosity of the investigated liquid to the viscosity of water, taken as a unit. To measure kinematic viscosity, various modifications of viscometers such as Ostwald and Ubbelohde are used. The kinematic viscosity is usually expressed in m 2 * s -1 . Knowing the density of the liquid under study, one can then calculate the dynamic viscosity, which is expressed in Pa * s. Dynamic viscosity can also be determined using rotational viscometers of various modifications such as "Polymer RPE-1 I" or microrheometers of the VIR series. Geppler-type viscometers are based on measuring the speed of a ball falling in a liquid. They allow you to set the dynamic viscosity. All instruments must be temperature controlled, as viscosity is highly dependent on the temperature of the fluid being tested.

Solubility in GF XI is considered not as a physical constant, but as a property that can serve as an approximate characteristic of the test preparation. Along with the melting point, the solubility of a substance at constant temperature and pressure is one of the parameters by which the authenticity and purity of almost all medicinal substances are established.

The method for determining the solubility according to SP XI is based on the fact that a sample of a pre-ground (if necessary) drug is added to a measured volume of the solvent and continuously mixed for 10 minutes at (20±2)°C. A drug is considered dissolved if no particles of the substance are observed in the solution in transmitted light. If the dissolution of the drug takes more than 10 minutes, then it is classified as slowly soluble. Their mixture with the solvent is heated on a water bath to 30°C and complete dissolution is observed after cooling to (20±2)°C and vigorous shaking for 1--2 minutes. More detailed instructions on the conditions for the dissolution of slowly soluble drugs, as well as drugs that form cloudy solutions, are given in private articles. Solubility rates in various solvents are indicated in private articles. They stipulate cases when solubility confirms the degree of purity of the medicinal substance.

In GF XI, no. 1 (p. 149) includes the phase solubility method, which makes it possible to quantify the degree of purity of a medicinal substance by accurately measuring solubility values. This method is based on the Gibbs phase rule, which establishes the relationship between the number of phases and the number of components under equilibrium conditions. The essence of establishing phase solubility lies in the successive addition of an increasing mass of the drug to a constant volume of the solvent. To achieve a state of equilibrium, the mixture is subjected to prolonged shaking at a constant temperature, and then, using diagrams, the content of the dissolved medicinal substance is determined, i.e. establish whether the test preparation is an individual substance or a mixture. The phase solubility method is characterized by objectivity, does not require expensive equipment, knowledge of the nature and structure of impurities. This makes it possible to use it for qualitative and quantitative analyses, as well as for studying the stability and obtaining purified drug samples (up to a purity of 99.5%). An important advantage of the method is the ability to distinguish between optical isomers and polymorphic forms of drugs. The method is applicable to all kinds of compounds that form true solutions.

2.2 Setting the pH of the medium

Important information about the degree of purity of the medicinal product is given by the pH value of its solution. This value can be used to judge the presence of impurities of acidic or alkaline products.

The principle of detecting impurities of free acids (inorganic and organic), free alkalis, i.e. acidity and alkalinity, is to neutralize these substances in a solution of the drug or in an aqueous extract. Neutralization is performed in the presence of indicators (phenolphthalein, methyl red, thymolphthalein, bromophenol blue, etc.). The acidity or alkalinity is judged either by the color of the indicator, or by its change, or the amount of titrated alkali or acid solution used for neutralization is established.

The reaction of the medium (pH) is a characteristic of the chemical properties of a substance. This is an important parameter that should be set when performing technological and analytical operations. The degree of acidity or basicity of solutions must be taken into account when performing drug purity and quantitation tests. The shelf life of medicinal substances, as well as the severity of their use, depend on the pH values ​​of solutions.

The pH value approximately (up to 0.3 units) can be determined using indicator paper or a universal indicator. Of the many ways to establish the pH value of the environment, GF XI recommends colorimetric and potentiometric methods.

The colorimetric method is very simple to implement. It is based on the property of indicators to change their color at certain ranges of pH values. To perform the tests, buffer solutions with a constant concentration of hydrogen ions are used, differing from each other by a pH value of 0.2. To a series of such solutions and to the test solution add the same amount (2-3 drops) of the indicator. According to the coincidence of color with one of the buffer solutions, the pH value of the medium of the test solution is judged.

In GF XI, no. 1 (p. 116) provides detailed information on the preparation of standard buffer solutions for various pH ranges: from 1.2 to 11.4. As reagents for this purpose, combinations of various ratios of solutions of potassium chloride, potassium hydrophthalate, monosubstituted potassium phosphate, boric acid, sodium tetraborate with hydrochloric acid or sodium hydroxide solution are used. Purified water used for the preparation of buffer solutions should have a pH of 5.8--7.0 and be free from carbon dioxide impurities.

The potentiometric method should be attributed to physicochemical (electrochemical) methods. The potentiometric determination of pH is based on the measurement of the electromotive force of an element composed of a standard electrode (with a known potential value) and an indicator electrode, the potential of which depends on the pH of the test solution. To establish the pH of the medium, potentiometers or pH meters of various brands are used. Their adjustment is carried out using buffer solutions. The potentiometric method for determining pH differs from the colorimetric method in higher accuracy. It has fewer limitations and can be used to determine pH in colored solutions, as well as in the presence of oxidizing and reducing agents.

In GF XI, no. 1 (p. 113) includes a table that lists the solutions of substances used as standard buffer solutions for testing pH meters. The data given in the table make it possible to establish the temperature dependence of the pH of these solutions.

2.3 Determination of transparency and turbidity of solutions

Transparency and degree of turbidity of the liquid according to SP X (p. 757) and SP XI, vol. 1 (p. 198) is established by comparing the test tubes of the test liquid with the same solvent or with standards in a vertical arrangement. A liquid is considered transparent if, when it is illuminated with an opaque electric lamp (power 40 W), on a black background, the presence of undissolved particles, except for single fibers, is not observed. According to GF X, standards are a suspension obtained from certain amounts of white clay. Standards for determining the degree of turbidity according to SP XI are suspensions in water from mixtures of certain amounts of hydrazine sulfate and hexamethylenetetramine. First prepare a 1% solution of hydrazine sulfate and a 10% solution of hexamethylenetetramine. By mixing equal volumes of these solutions, a reference standard is obtained.

In the general article of SP XI, there is a table that indicates the quantities of the main standard required for the preparation of reference solutions I, II, III, IV. It also shows the scheme for viewing the transparency and degree of turbidity of liquids.

Coloring of liquids according to GF XI, vol. 1 (p. 194) is determined by comparing the test solutions with an equal amount of one of the seven standards in daylight reflected light on a matte white background. For the preparation of standards, four basic solutions are used, obtained by mixing in various ratios of the initial solutions of cobalt chloride, potassium dichromate, copper (II) sulfate and iron (III) chloride. Sulfuric acid solution (0.1 mol/l) is used as a solvent for the preparation of stock solutions and standards.

Liquids are considered colorless if they do not differ in color from water, and solutions - from the corresponding solvent.

Adsorption capacity and dispersion are also indicators of the purity of some drugs.

Very often, a test based on their interaction with concentrated sulfuric acid is used to detect impurities of organic substances. The latter can act as an oxidizing or dehydrating agent.

As a result of such reactions, colored products are formed. The intensity of the resulting color should not exceed the corresponding color standard.

To establish the purity of drugs, the definition of ash is widely used (GF XI, issue 2, p. 24). By calcining a sample of the preparation in a porcelain (platinum) crucible, the total ash is established. Then, after adding diluted hydrochloric acid, the ash insoluble in hydrochloric acid is determined. In addition, sulfate ash obtained after heating and calcining a sample of the preparation treated with concentrated sulfuric acid is also determined.

One of the indicators of the purity of organic drugs is the content of the residue after calcination.

When establishing the purity of some drugs, they also check the presence of reducing substances (by discoloration of the potassium permanganate solution), coloring substances (colorlessness of the aqueous extract). Water-soluble salts (in insoluble preparations), substances insoluble in ethanol, and impurities insoluble in water (according to the turbidity standard) are also detected.

2.4 Estimation of chemical constants

To assess the purity of oils, fats, waxes, and some esters, chemical constants such as acid number, saponification number, ester number, iodine number are used (SP XI, issue 1, pp. 191, 192, 193).

Acid number - the mass of potassium hydroxide (mg), which is necessary to neutralize the free acids contained in 1 g of the test substance.

Saponification number - the mass of potassium hydroxide (mg), which is necessary to neutralize free acids and acids formed during the complete hydrolysis of esters contained in 1 g of the test substance.

The ester number is the mass of potassium hydroxide (mg) that is needed to neutralize the acids formed during the hydrolysis of esters contained in 1 g of the test substance (i.e. the difference between the saponification number and the acid number).

The iodine number is the mass of iodine (g) that binds 100 g of the test substance.

SP XI provides methods for establishing these constants and methods for calculating them.

Chapter 3. Chemical Methods of Analysis

3.1 Features of chemical methods of analysis

These methods are used to authenticate medicinal substances, test them for purity, and quantify them.

For identification purposes, reactions are used that are accompanied by an external effect, such as a change in the color of the solution, the release of gaseous products, precipitation or dissolution of precipitates. Establishing the authenticity of inorganic medicinal substances consists in detecting, using chemical reactions, the cations and anions that make up the molecules. Chemical reactions used to identify organic medicinal substances are based on the use of functional analysis.

The purity of medicinal substances is established by means of sensitive and specific reactions, suitable for determining the acceptable limits for the content of impurities.

Chemical methods have proved to be the most reliable and effective, they allow you to perform the analysis quickly and with high reliability. In case of doubt in the results of the analysis, the last word remains with the chemical methods.

Quantitative methods of chemical analysis are divided into gravimetric, titrimetric, gasometric analysis and quantitative elemental analysis.

3.2 Gravimetric (weight) method

The gravimetric method is based on the weighing of the precipitated substance in the form of a poorly soluble compound or the distillation of organic solvents after the extraction of the medicinal substance. The method is accurate but lengthy, as it involves such operations as filtering, washing, drying (or calcining) to constant weight.

Sulphates can be determined gravimetrically from inorganic medicinal substances by converting them into insoluble barium salts, and silicates by preliminary calcination to silicon dioxide.

Methods for gravimetric analysis of preparations of quinine salts recommended by the Global Fund are based on the precipitation of the base of this alkaloid under the action of sodium hydroxide solution. Bigumal is determined in the same way. Benzylpenicillin preparations are precipitated as N-ethylpiperidine salt of benzylpenicillin; progesterone - in the form of hydrazone. It is possible to use gravimetry to determine alkaloids (by weighing free bases or picrates, picrolonates, silicotungstates, tetraphenylborates), as well as to determine some vitamins that are precipitated in the form of water-insoluble hydrolysis products (vikasol, rutin) or in the form of silicotungstate (thiamine bromide ). There are also gravimetric techniques based on the precipitation of acidic forms of barbiturates from sodium salts.

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Page 1

One of the most important tasks of pharmaceutical chemistry is the development and improvement of methods for assessing the quality of medicines.

To establish the purity of medicinal substances, various physical, physico-chemical, chemical methods of analysis or a combination of them are used. GF offers the following methods of drug quality control.

Physical and physico-chemical methods. These include: determination of melting and solidification temperatures, as well as temperature limits of distillation; determination of density, refractive indices (refractometry), optical rotation (polarimetry); spectrophotometry - ultraviolet, infrared; photocolorimetry, emission and atomic absorption spectrometry, fluorimetry, nuclear magnetic resonance spectroscopy, mass spectrometry; chromatography - adsorption, distribution, ion-exchange, gas, high-performance liquid; electrophoresis (frontal, zonal, capillary); electrometric methods (potentiometric determination of pH, potentiometric titration, amperometric titration, voltammetry).

In addition, it is possible to use methods that are alternative to pharmacopoeial methods, which sometimes have more advanced analytical characteristics (speed, accuracy of analysis, automation). In some cases, a pharmaceutical company purchases a device based on a method not yet included in the Pharmacopoeia (for example, the Raman spectroscopy method - optical dichroism). Sometimes it is advisable to replace the chromatographic method with a spectrophotometric one when determining the authenticity or testing for purity. The pharmacopoeial method for determining heavy metal impurities by precipitating them in the form of sulfides or thioacetamides has a number of disadvantages. To determine heavy metal impurities, many manufacturers are implementing physicochemical methods of analysis such as atomic absorption spectrometry and inductively coupled plasma atomic emission spectrometry.

An important physical constant that characterizes the authenticity and degree of purity of drugs is the melting point. A pure substance has a distinct melting point, which changes in the presence of impurities. For medicinal substances containing a certain amount of admissible impurities, the GF regulates the melting temperature range within 2 °C. But in accordance with Raoult's law (AT = iK3C, where AT is the decrease in the crystallization temperature; K3 is the cryoscopic constant; C is the concentration) at i = 1 (non-electrolyte), the value of AT cannot be the same for all substances. This is connected not only with the content of impurities, but also with the nature of the drug itself, i.e., with the value of the cryoscopic constant K3, which reflects the molar decrease in the melting point of the drug. Thus, at the same AT = 2 "C for camphor (K3 = 40) and phenol (K3 = 7.3), the mass fractions of impurities are not equal and amount to 0.76 and 2.5%, respectively.

For substances that melt with decomposition, the temperature at which the substance decomposes and a sharp change in its appearance occurs is usually indicated.

Purity criteria are also the color of the drug and / or the transparency of liquid dosage forms.

Physical constants such as the refractive index of a light beam in a solution of the test substance (refractometry) and the specific rotation due to the ability of a number of substances or their solutions to rotate the polarization plane when gaussically polarized light passes through them (polarimetry) can serve as a certain criterion for the purity of a drug. Methods for determining these constants are related to optical methods of analysis and are also used to establish the authenticity and quantitative analysis of drugs and their dosage forms.

An important criterion for the good quality of a number of drugs is their water content. A change in this indicator (especially during storage) can change the concentration of the active substance, and, consequently, the pharmacological activity and make the drug unsuitable for use.

Chemical methods. These include: qualitative reactions for authenticity, solubility, determination of volatile substances and water, determination of nitrogen content in organic compounds, titrimetric methods (acid-base titration, titration in non-aqueous solvents, complexometry), nitritemetry, acid number, saponification number, ether number, iodine number, etc.

biological methods. Biological methods of drug quality control are very diverse. Among them are tests for toxicity, sterility, microbiological purity.

UDC 615.015:615.07:53

ANALYSIS OF DRUGS FOR PHARMACOKINETIC

RESEARCH

Dmitry Vladimirovich Reyhart1, Viktor Vladimirovich Chistyakov2

Department of Organization and Management in the Sphere of Medicines Circulation (Head - Corresponding Member of the Russian Academy of Medical Sciences, Prof. R.U. Khabriev) of the Moscow State Medical Academy. THEM. Sechenov,

2 Center for Chemistry of Medicinal Products - VNIHFI (general director - K.V. Shilin), Moscow

A review of sensitive and specific analytical methods used in the study of the pharmacokinetics of drugs was carried out. The advantages and limitations of the use of enzyme immunoassay, high performance liquid chromatography with fluorescent and mass spectrometric detection are shown. The use of one or another method in assessing the pharmacokinetics of drugs in each case is determined by the structure of the test compound and the equipment of the laboratory.

Key words: liquid chromatography, fluorescence and mass spectrometric detection, enzyme immunoassay, pharmacokinetics.

The study of pharmacokinetics is based mainly on the assessment of the concentration in the patient's body of a drug substance (PM) at certain points in time after taking the drug. The object of the study is blood (whole, serum, plasma), urine, saliva, feces, bile, amniotic fluid, etc. The most accessible and most frequently studied blood and urine samples.

Measuring the concentration of a drug can be divided into two stages: 1 - isolation of a specific drug substance from a biological object, concentration of the test compound, separating it from the main endogenous components; 2 - separation of a mixture of compounds, identification of drugs and quantitative analysis.

The study of the concentration of the drug in the blood provides information on the duration of circulation of the drug in the body, the bioavailability of the drug, the effect of concentration on the pharmacological effect, therapeutic and lethal doses, the dynamics of the formation of active or toxic metabolites.

The study of the concentration of the drug in the urine allows you to evaluate the rate of elimination of drugs and kidney function. The concentration of metabolites in the urine is an indirect indicator of the activity of metabolizing enzymes.

The study of biological material includes measuring the mass (volume) of the sample, the release of the drug (metabolites) from 532

sample cells, separation of whole cells (for example, when analyzing blood) or parts of cells (when analyzing tissue homogenates), adding an internal standard, separating proteins, sample purification (centrifugation, filtration), extraction, re-extraction, concentration and conversion of test substances into convenient for the analysis of derivatives, the main procedures for processing blood and urine samples, respectively (Fig. 1).

The “ideal” analytical method for measuring drug concentration should have high sensitivity, specificity and reproducibility, the ability to work with small volumes, ease of material preparation, low cost and ease of equipment maintenance, reliability and automation, ease of personnel work and versatility (the ability to analyze various classes of drugs) .

To obtain reliable data, it is necessary to correct for the stability of the active substance and / or product (s), as well as the degree of its biotransformation in the analyzed biological media.

The validation of a method should be based on its intended application, and the calibration should take into account the concentration range of the test sample. It is strongly discouraged to use two or more methods of sample analysis on the same material with a similar range of calibration values.

There are a large number of methods for determining the concentration of drugs in biological fluids: chromatographic, microbiological, spectrophotometric, polarographic, immunological (radioimmune, immunoenzymatic), radioisotope and other methods.

The critical parameters of the method are sensitivity, speed, accuracy, the ability to work with a small amount of biomaterial and cost.

In table. 1 compares analytical methods for drug analysis.

The most widely (up to 95% of studies) in practice is the method of highly effective

Rice. 1. Basic procedures for handling blood and urine samples.

noah liquid chromatography (HPLC) with various types of detection.

The advantages of HPLC compared, for example, with gas-liquid chromatography (GLC) are the absence of restrictions on the thermal stability of the analyzed preparations, the ability to work with aqueous solutions and volatile compounds, and the use of “normal-phase” and “reverse-phase” chromatography options. Many of the types of detection are non-destructive.

enzyme immunoassay, HPLC with fluorescent detection, HPLC with mass spectrometric detection, which are currently actively used in pharmacokinetic studies.

ELISA method

The method of enzyme immunoassay (ELISA) was proposed in the early 70s of the last century. The principle of ELISA is the interaction of specific protein proteins

Comparative characteristics of drug analysis methods

Methods Absolute sensitivity, g Sensitivity, points Complexity, points Selectivity, points Universality Total assessment, points

Liquid chromatography:

UV detector 10-7 3 -3 4 4 8

fluorescent detector 10-8 - 10-9 4 -3 5 2 8

mass spectrometric detector 10-11 - 10-12 5 -5 5 4 9

Immunological 10-10 - 10-11 5 -1 4 1 9

Gas Chromatography:

electron capture detector 10-10 5 -4 4 2 7

flame ionization detector 10-8 - 10-9 4 -3 2 4 7

mi; detection methods used in HPLC have higher specificity.

Let us consider the features of highly sensitive methods that allow us to analyze nanogram amounts of drugs (Table 1):

a body with an analyte acting as an antigen. The higher the concentration of the substance-antigen, the more antigen-antibody complexes are formed. For the quantitative analysis of complex formation at-

two approaches change - with preliminary separation of the complex (heterogeneous methods) or without its separation (homogeneous methods). In both cases, a sample with an unknown analyte concentration is added to serum in which the antibody is complexed with a labeled analogue of the analyte, and the substance from the analyzed sample is displaced from the complex. The amount of displaced labeled analogue is proportional to the concentration of the substance in the sample. Having determined how much of the labeled analog turned out to be displaced from the complex (or, on the contrary, remained bound), it is possible to calculate the desired level of the substance in the sample. Pre-calibration is carried out using standard solutions (with standard concentrations of the test substance).

Reagent kits are produced - the so-called diagnosticums (antiserum, enzyme, substrate, cofactor, standard solutions for calibration combined with the drug), designed for 50-200 tests. For analysis, 0.05-0.2 ml of the patient's blood serum is usually sufficient.

Immunoenzyme methods have high sensitivity and specificity. Diagnostic kits are relatively cheap and have a longer shelf life than kits for radioimmunoassays. When using ELISA, the need to separate the antigen-antibody complex is eliminated - a rather complicated procedure, with a relatively high risk of error. The immunoenzymatic method can be performed in any hospital or outpatient laboratory; devices have been developed that provide full automation of analysis.

Ease of analysis, high sensitivity, accuracy, reproducibility,

the moderate price of equipment and reagents - all this creates the prospect for the widespread introduction of immunological methods into medical practice.

High performance liquid chromatography with fluorescence detection

In HPLC, the detector generates an electrical signal whose strength is proportional to the concentration of the analyte dissolved in the mobile phase. In the first liquid chromatographs (ion-exchange), the mobile phase passing through the column with the sample components was collected in small vessels, and then using titrometry, colorimetry, polarography, etc. the content of the component in this portion was determined. In other words, sample separation processes

and definitions of its quantitative composition were separated in time and space. In a modern liquid chromatograph, these processes are provided by one instrument.

Any physicochemical property of the mobile phase (absorption or emission of light, electrical conductivity, refractive index, etc.) that changes in the presence of molecules of separable compounds can be used to detect sample components. Of the existing 50 physicochemical detection methods, 5-6 are currently actively used.

Sensitivity is the most important characteristic of the detector. If the sensitivity is determined in terms of the double amplitude of the noise of the zero line, and the noise is expressed in physical units, then the sensitivity of the photometric detector will be expressed in units of optical density, the sensitivity of the refractometric detector in units of the refractive index, the voltammetric detector in amperes, and the conductometric detector in siemens. In pharmaceutical analysis, sensitivity is expressed in terms of the minimum amount of analyte. The degree of sensitivity of various types of detectors is given in Table. one.

Despite the fact that currently 80% of chromatographs are equipped as standard with spectrophotometric detectors, fluorescence detection is becoming more widespread, especially when determining the concentration of compounds that can “glow” under the action of exciting radiation. The luminescence intensity is proportional to the intensity of the exciting light. The study of emission spectra (fluorescence and phosphorescence) is a more sensitive and specific method than the study of absorption spectra.

The fluorescence spectrum of a substance in many cases is a mirror image of the absorption band with the lowest energy and is usually located next to this band on its long wavelength side. This method is most conveniently used in the study of drugs that have their own fluorescence (chloroquine, doxorubicin, doxazosin, atenolol, indomethacin, propranolol, tetracyclines, quinidine, etc.). Some drugs can be converted relatively easily into fluorescent compounds (derivatization process), such as hydrocortisone (treatment with sulfuric acid), meperidine (condensation with formaldehyde), 6-mercap-topurine, and methotrexate (oxidation with potassium permanganate). Other drugs with active functional groups can be condensed with fluorescent reagents.

gentami - fluorescamine (chlordeazepoxide, novocainamide, sulfonamides, etc.), 7-nitrobenzo-2,1,3-oxadiazole (propoxyphene, etc.), etc. At the same time, it should be noted that, with high sensitivity and selectivity, fluorescent detection methods are limited by the range of drugs that have natural fluorescence, and the derivatization process in quantitative analysis is costly.

High Performance Liquid Chromatography with Mass Spectrometric Detection

A highly sensitive version of the modern HPLC detector used for pharmacokinetic studies is the mass spectrometer. The mass spectrometric detector can significantly reduce the analysis time, in particular, by eliminating the preparatory stage (extraction). This method makes it possible to simultaneously identify several substances, and this eliminates errors associated with the presence of inseparable components.

Mass spectrometry is one of the most promising methods for the physicochemical analysis of drugs. Traditionally, organic mass spectrometry is used to solve two main problems: the identification of substances and the study of the fragmentation of ionized molecules in the gas phase. The combination of a mass spectrometer with a liquid chromatograph significantly expanded the possibilities of the classical method. With the advent of new ionization methods, such as "electrospray" (ESI - English electrospray ionization) - ionization in an electric field at atmospheric pressure) and "MALDI" - ionization by laser desorption, the list of molecules that can be studied by this method has expanded significantly.

Currently, the combination of HPLC and an "electrospray" mass spectrometric detector is widely used in pharmacokinetic and bioequivalence studies of drugs. Initially, the ESI method was developed under the leadership of L.N. Gall, and in 2002 D. Fennu and K. Tanaka were awarded the Nobel Prize for the development of methods for the identification and structural analysis of biological macromolecules and, in particular, methods for the mass spectrometric analysis of biological macromolecules. There are three stages in the mechanism of formation of ionized particles. The first is the formation of charged droplets at the capillary section. Through the applied voltage, the charge is redistributed in the solution, the positive ions are trapped

spilling out at the exit. With a strong applied field (3-5 kV), a jet is formed from the top of the cone, which then scatters into small drops. The second stage is the gradual reduction in the size of the charged droplets due to the evaporation of the solvent and the subsequent disintegration of the droplets up to the formation of true ions. The charged droplets move through the atmosphere towards the opposite electrode. The third stage is repeated cycles of separation and reduction in the volume of droplets until the solvent is completely evaporated and ions are formed in the gas phase.

Modern LC/MS systems (LC/MS - liquid chromatography/mass-spectrometry) make it possible to register the total ion current (TIC - total ion current), control the specified ions (SIM - selected ion monitoring) and control the specified reactions selective reaction monitoring (SRM - English selected reaction monitoring).

Total ionic current (TIC) analysis provides data on all compounds sequentially exiting the chromatographic column. Mass chromatograms resemble chromatograms with UV detection, with the area under the peak corresponding to the amount of substance. When determining target ions (SIM), the operator can limit the detection range of the required compounds by isolating, for example, minor substances. The SRM method has the highest sensitivity and specificity when the ion current is recorded using one selected ion characteristic of the compound under study (in ESI ionization and registration of positive ions, this is usually the MH+ molecular ion).

Recently published papers discuss the possibility of quantitative analysis of organic substances in biological objects without chromatographic separation using multi-on detection and internal control in the form of a deuterium-labeled analog. In particular, for molecules of a lipid nature, the concentration range (from pico- to nanomolar) was determined, at which the authors observed a linear dependence of the intensity of the ion current on the concentration of the substance. An increase in the concentration of compounds in the solution led to ion-molecular interactions during ionization and violation of linearity.

A method for the quantitative determination of prostaglandins and polyunsaturated fatty acids using electrospray ionization - mass spectrometry without chromatographic separation using an internal standard and registration of negative ions is described. In work

Yu.O. Caratasso and I. V. Logunova, the sensitivity of mass spectrometry in the study of a potential antiarrhythmic agent was 3 ng/0.5 ml of blood plasma.

When choosing an analytical method, it must be borne in mind that the use of ELISA is limited by the presence of required reagents, fluorescent detection, and the need for intrinsic fluorescence in the test compound. Although the above limitations are not significant for mass spectrometric detection, the cost of equipment today remains quite high, and this type of analysis requires special skills.

LITERATURE

1. Aleksandrov M.L., Gall L.N., Krasnov N.V. Ion extraction from solutions at atmospheric pressure - a new method of mass spectrometric analysis // Dokl. Acad. sciences of the USSR. - 1984. - T.277. - No. 2. -

2. Karatasso Yu.O., Logunova IV, Sergeeva MG et al. Quantitative analysis of drugs in blood plasma using electrospray ionization-mass spectrometry without chromatographic separation // Khim. farm. magazine - 2007. - No. 4. - S. 161-166.

3. Karatasso Yu.O., Alyoshin S.E., Popova N.V. Quantitative analysis of prostaglandins and polyunsaturated fatty acids by mass spectrometry with electrospray ionization // Mass spectrometry. -2007. - T.4. - IN 3. - S. 173-178.

4. Kholodov L.E., Yakovlev V.P. Clinical pharmacokinetics. - M.: Medicine, 1985. - 463 p.

5. Covey T.R., Lee E.D., Henion J.D. High-speed liquid chromatography/tandem mass spectrometry for the determination of drugs in biological samples // Anal. Chem. - 1986. - Vol. 58 (12). - P. 2453-2460.

6. Conference report on analytical methods validation: bioavailability, bioequivalence and pharmacokinetic studies // J. Pharmac. sci. - 1992. - Vol.81. - P. 309-312.

7. De Long C.J., Baker P.R.S., SamuelM. et al. Molecular species composition of rat liver phospholipids by ESI-MS/ MS: The effect of chromatography//J. Lipid Res. - 2001. - Vol. 42. - P. 1959-1968.

8. Electrospray Ionization Mass Spectrometry. Ed. R. B. Cole // Wiley. - New York, 1997.

9. Han X., Yang K., Yang J. et al. Factors influencing the electrospray intrasource separation and selective ionization of glycerophospholipids // Am. soc. mass spectrom. - 2006. - Vol. 17(2). - P. 264-274.

10. Koivusalo M., Haimi P., Heikinheimo L. et al. Quantitative determination of phospholipids compositions by ESI-MS: Effects of acyl chain length, unsaturation, and lipid concentration on instrument response // J. Lipid Res. - 2001. - Vol. 42.-P. 663-672.

11. Lee M.S., Kerns E.H. LC/MS applications in drug discovery//Mass Spectrom. Rev. - 1999. - Vol. 18(3-4). - P. 187-279.

Received 05/28/10.

ANALYSIS OF DRUGS IN PHARMACOKINETIC STUDIES

D.V. Reikhart, V.V. Chistyakov

Conducted was a review of sensitive and specific analytical methods for studying the pharmacokinetics of drugs. Shown were the advantages and limitations of immune-enzyme analysis, of high performance liquid chromatography with fluorescence and mass spectrometric detection. The usage of a method in the evaluation of the pharmacokinetics of drugs in each case should be determined by the structure of the compound and the laboratory equipment.

Key words: liquid chromatography, fluorescence and mass spectrometric detection, immune-enzyme analysis, pharmacokinetics.

MINISTRY OF EDUCATION

STATE BUDGET EDUCATIONAL INSTITUTION OF HIGHER PROFESSIONAL EDUCATION "SIBERIAN

STATE MEDICAL UNIVERSITY" OF THE MINISTRY OF HEALTH AND SOCIAL DEVELOPMENT OF THE RUSSIAN FEDERATION

Analysis of complex dosage forms

Part 1. Dosage forms of pharmaceutical production

Tutorial

For self-training and a guide to laboratory classes in pharmaceutical chemistry for students of pharmaceutical faculties of universities of full-time and part-time education

UDC 615.07 (071) BBK R 282 E 732

E.V. Ermilova, V.V. Dudko, T.V. Kadyrov Analysis of complex dosage forms Part 1. Pharmaceutical production dosage forms: Uch. allowance. - Tomsk: Ed. 20012 . – 169 p.

The manual contains methods for the analysis of dosage forms of pharmaceutical production. It discusses the terminology, classification of dosage forms, provides regulatory documents that control the quality of medicines in pharmacy production, indicates the features of intra-pharmacy express analysis; the main stages of the analysis of dosage forms are described in detail, while special attention is paid to chemical control.

The main part of the manual is devoted to the presentation of material on the analysis of dosage forms: liquid (mixtures, sterile) and solid (powders), numerous examples are given.

The appendix contains extracts from orders, refractometric tables, information on indicators, forms of reporting journals.

For students of pharmaceutical faculties of higher educational institutions.

Tab. 21. Fig. 27. Bibliography: 18 titles.

Preface. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

I. INTRODUCTION TO DOSAGE ANALYSIS

1.1. Terms used in pharmacy. . . . . . . . . . . . . . . . ………. 5 1.1.1. Terms characterizing medicines.. ….5 1.1.2. Terms characterizing dosage forms. . . ….5 1.2. Classification of dosage forms. . . . . . . . . . . . . . . . . . . . . . 7

1.3. Normative documents and requirements for the quality of medicines of pharmaceutical production. . . . . . . . . . . . . …...7 1.4. Peculiarities of express-analysis of medicinal products of pharmaceutical production. . . . . . . . . . . . . . . . . . . . . . . . . . ……………8

1.4.1. Features of determining the authenticity of the express method. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ………..nine

1.4.2. Features of quantitative express analysis. . . . . . . . …nine

2.1. Organoleptic and physical control. . . . . . . . . . . . . . . . . . 10 2.1.1. Organoleptic control. . . . . . . . . . . . . . . . . . . . . . . . . . .10 2.1.2. Physical control. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .10 2.2. Chemical control. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .11 2.2.1. Tests for authenticity. . . . . . . . . . . . . . . . . . . . . . . . . . . . .11 2.2.2.. Quantitative analysis. . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . fourteen

2.2.2.1. Ways of expressing concentrations. . . . . . . . . . . . . . . . .15 2.2.2.2. Methods of titrimetric analysis. . . . . . . . . . . . . . . 16 2.2.2.3. Calculation of the mass (volume) of the dosage form and the volume of the titrant for analysis. . . . . . . . . . . . . . . . . . . . . 17

2.2.2.4. Processing of measurement results. . . . . . . . . . . . . . . . . .19 2.2.2.5. Formulation of analysis results. . . . . . . . . . . . . . . . . . 32

III. ANALYSIS OF DOSAGE FORMS

Liquid dosage forms. . . . . . . . . . . . . . . . . . . . . . . . . . . . .33

3.1. Mixture analysis. . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . .33 3.2. Analysis of sterile dosage forms. . . . . . . . . . . . . . . . . . . . .59

Solid dosage forms

3.3. Powders. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .89

Questions of self-training control. . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Test control. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .125

Test control responses. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .130

APPLICATIONS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .131

Bibliography. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .168

Foreword

The basis for writing the textbook was the program in pharmaceutical chemistry for students of pharmaceutical universities (faculties)

M.: GOU VUNMTS, 2003

One of the components of pharmaceutical analysis is the analysis of pharmacy and factory-produced drugs, carried out by the methods of pharmacopoeial analysis, according to the requirements of various guidelines,

manuals, instructions, etc.

The manual is devoted to the methods of research of dosage forms

(potions, sterile, powders) manufactured in a pharmacy, where all types of intra-pharmacy control are used, but the most effective is chemical control, which makes it possible to check the compliance of the manufactured dosage form with the prescription, both in terms of authenticity and quantitative content. Authenticity and quantitation procedures are presented in such a way as to use the best methods of investigation, and the minimum amount of drug was spent on the analysis.

The main part contains numerous examples of the use of refractometry in the quantitative analysis of drugs, since this method is widely used in pharmacy practice.

The proposed textbook contributes to the development of students' chemical analytical thinking.

I. INTRODUCTION TO DOSAGE ANALYSIS

1.1. Terms used in pharmacy

1.1.1. Terms characterizing medicines

Medicines - substances used for prevention

diagnosis, treatment of disease, prevention of pregnancy, derived from

biological technologies.

medicinal substance- a medicinal product, which is an individual chemical compound or biological substance.

medicinal product- a medicinal product in the form of a specific

dosage form.

Dosage form- a condition that is convenient for use in which the desired therapeutic effect is achieved is attached to a medicinal product or medicinal plant material.

1.1.2. Terms characterizing dosage forms

Powders are a solid dosage form for internal and external use, consisting of one or more crushed substances and having the property of flowability.

Tablets - a dosage form obtained by pressing drugs or a mixture of drugs and excipients, intended for internal, external, sublingual,

implantation or parenteral use.

Capsules - a dosage form consisting of a drug enclosed in a shell.

Ointments are a soft dosage form intended for application to the skin, wounds or mucous membranes and consisting of a medicinal substance and a base.

Pastes - ointments with a content of powdery substances over 20-25%.

Suppositories are a dosage form that is solid at room temperature and melts at body temperature.

Solutions liquid dosage form obtained by dissolving one or more medicinal substances intended for injection, internal or external use.

Drops liquid dosage form intended for internal or external use, dosed in drops.

Suspensions are a liquid dosage form containing, as a dispersed phase, one or more powdered medicinal substances distributed in a liquid dispersion medium.

Emulsions uniform in appearance dosage form,

consisting of mutually insoluble finely dispersed liquids,

intended for internal, external or parenteral use.

Extracts - concentrated extracts from medicinal plant materials. There are liquid extracts (Extracta fluida); thick extracts (Extracta spissa) - viscous masses with a moisture content of not more than 25%;

dry extracts (Extracta sicca) - free-flowing masses with a moisture content of not more than

Infusions dosage form, which is an aqueous extract from medicinal plant materials or an aqueous solution of dry or liquid extracts (concentrates).

Decoctions infusions that differ in the mode of extraction.

Aerosols dosage form in which drugs and excipients are under the pressure of a propellant gas

(propellant) in an aerosol can, hermetically sealed with a valve.

1.2. Classification of dosage forms

Classification of dosage forms is carried out depending on:

1.2.1. Aggregate state Solid : powders, tablets, dragees, granules, etc.

Liquid: true and colloidal solutions, drops, suspensions, emulsions,

liniments, etc.

Soft: ointments, suppositories, pills, capsules, etc.

Gaseous: aerosols, gases.

1.2.2. Quantities of medicinal substances

One-component

Multicomponent

1.2.3. Places of manufacture

Factory

Pharmacy

1.2.4. Manufacturing method

Solutions for injections Medicines Eye drops Decoctions Infusions Aerosols Infusions

Homeopathic remedies, etc.

1.3. Regulatory documents and quality requirements

medicines of pharmaceutical production

All production activities of the pharmacy should be aimed at ensuring high-quality manufacturing of medicines.

One of the most important factors determining the quality of medicines manufactured in a pharmacy is the organization of intra-pharmacy control.

Intra-pharmacy control is a set of measures aimed at timely detection and prevention of errors that occur in the process of manufacturing, processing and dispensing medicines.

Pharmaceutical production drugs are subject to several types of control, depending on the nature of the dosage form.

The system of intra-pharmacy quality control of medicinal products provides for preventive measures, acceptance, organoleptic, written, questionnaire, physical, chemical and dispensing control.

According to the instructions of the Ministry of Health of the Russian Federation "On quality control of medicines manufactured in pharmacies" (Order No. 214 dated July 16, 1997), all medicines are subject to intra-pharmacy control: organoleptic, written and dispensing control - mandatory, questionnaire and physical - selectively, and chemical - in accordance with paragraph 8 of this order (see Appendix).

1.4. Features of express analysis of medicines

pharmacy production

The need for intra-pharmacy control is due to the corresponding high quality requirements for medicines manufactured in pharmacies.

Since the manufacture and distribution of drugs in pharmacies is limited to a short time, their quality is assessed by express methods.

The main requirements for express analysis are the consumption of minimal quantities of drugs with sufficient accuracy and sensitivity, simplicity and speed of execution, if possible, without separation of ingredients, the possibility of conducting an analysis without removing the prepared medicinal product.

If it is not possible to perform the analysis without separating the components, then the same separation principles are used as in macro analysis.

1.4.1. Features of determining the authenticity of the express method

The main difference between determining the authenticity of the express method from macro-analysis is the use of small amounts of the studied mixtures without separating them.

The analysis is performed by the drip method in micro-test tubes, porcelain cups, on watch glasses, while 0.001 to 0.01 g of powder or 15 drops of the test liquid are consumed.

To simplify the analysis, it is sufficient to carry out one reaction for a substance, and the simplest, for example, for atropine sulfate, it is enough to confirm the presence of a sulfate ion, for papaverine hydrochloride - a chloride ion by classical methods.

1.4.2. Features of quantitative express analysis

Quantitative analysis can be performed by titrimetric or physico-chemical methods.

Titrimetric express analysis differs from macro methods in the consumption of smaller quantities of analyzed preparations: 0.05 0.1 g of powder or 0.5 2 ml of solution, and the exact mass of the powder can be weighed on hand-held scales; to improve accuracy, dilute solutions of titrants can be used: 0.01 0.02 mol/l.

A weighed portion of a powder or a volume of a liquid dosage form is taken in such a way that 1–3 ml of the titrant solution is used for the determination.

Of the physicochemical methods in pharmacy practice, the economical method of refractometry is widely used in the analysis of concentrates,

semi-finished products and other dosage forms.

II. MAIN STAGES OF PHARMACEUTICAL ANALYSIS

2.1. Organoleptic and physical control

2.1.1. Organoleptic control

Organoleptic control consists in checking the dosage form for the following indicators: appearance (“Description”), smell,

homogeneity, absence of mechanical impurities. The taste is checked selectively, and dosage forms prepared for children - everything.

Uniformity of powders, homeopathic triturations, ointments, pills,

suppositories are checked before dividing the mass into doses in accordance with the requirements of the current State Pharmacopoeia. The check is carried out selectively at each pharmacist during the working day, taking into account the types of dosage forms. The results of organoleptic control are recorded in the journal.

2.1.2. Physical control

Physical control consists in checking the total mass or volume of the dosage form, the number and mass of individual doses (at least three doses),

included in this dosage form.

This checks:

Each series of packaging or intra-pharmaceutical blanks in the amount of at least three packages;

Dosage forms manufactured according to individual prescriptions (requirements), selectively during the working day, taking into account all types of dosage forms, but not less than 3% of the number of dosage forms manufactured per day;