Chemical methods of drug research. Methods of drug analysis

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

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 obtained 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 the minimum 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, criteria such 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 can be detected by this method with a given confidence level. 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 allow determining 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 Errors in 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 ±(3-3.5) %; 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 one. 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 stages. 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 related 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 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

8-09-2015, 20:00

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The biological assessment of the quality of drugs is usually carried out according to the strength of the pharmacological effect or toxicity. Biological methods are used when physical, chemical or physico-chemical methods fail to make a conclusion about the purity or toxicity of the medicinal product, or when the method of preparation of the drug does not guarantee the constancy of activity (for example, antibiotics).

Biological tests are carried out on animals (cats, dogs, rabbits, frogs, etc.), individual isolated organs (uterine horn, part of the skin), individual groups of cells (blood cells), as well as on certain strains of microorganisms. The activity of drugs is expressed in units of action (ED).

Biological control of drugs containing cardiac glycosides. According to SP XI, a biological assessment of the activity of medicinal plant materials and preparations derived from it containing cardiac glycosides, in particular foxglove (purple, large-flowered and woolly), adonis, lily of the valley, strophanthus, gray jaundice, is carried out. Tests are carried out on frogs, cats and pigeons, setting the frog (ICE), feline (CED) and pigeon (CED) action units, respectively. One ICE corresponds to the dose of the standard sample, which, under experimental conditions, causes systolic cardiac arrest in the majority of experimental standard frogs (males weighing 28–33 g). One KED or GED corresponds to the dose of a standard sample or test drug per 1 kg of animal or bird weight that causes systolic cardiac arrest in a cat or pigeon. The ED content is calculated in 1.0 g of the study drug, if plant materials or dry concentrates are tested; in one tablet or in 1 ml if liquid dosage forms are being tested.

Toxicity test. In this section GF XI, no. 2 (p. 182), in comparison with SP X, a number of additions and changes have been made, reflecting the increasing requirements for the quality of medicines and the need to unify the conditions for their testing. The article includes a section that describes the procedure for sampling. The mass of animals on which the test is carried out has been increased, the conditions for their maintenance and the period of observation of them have been indicated. To perform the test, two vials or ampoules are selected from each batch containing not more than 10,000 vials or ampoules. From parties with a large number, three ampoules (vials) are selected from each series. The contents of samples of one series are mixed and tested on healthy white mice of both sexes weighing 19–21 g. The test solution is injected into the tail vein of five mice and animals are observed for 48 hours. The drug is considered to have passed the test if none of the experimental mice die in within the specified period. In the event of the death of even one mouse, the test is repeated according to a certain scheme. Private articles may also specify a different procedure for conducting a toxicity test.

Pyrogenicity tests. Bacterial pyrogens are substances of microbial origin that can cause in humans and warm-blooded animals when they enter the bloodstream channel fever, leukopenia, drop in blood pressure and other changes in various organs and systems of the body. The pyrogenic reaction is caused by gram-negative living and dead microorganisms, as well as their decay products. Permissible content, for example, in an isotonic solution of sodium chloride, 10 microorganisms per 1 ml, and with the introduction of not more than 100 ml, 100 per 1 ml is allowed. The test for pyrogenicity is subjected to water for injection, injection solutions, immunobiological drugs, solvents used for the preparation of injection solutions, as well as dosage forms that cause, according to clinics, a pyrogenic reaction.

In SP XI, as well as in the pharmacopoeias of other countries of the world, a biological method for testing pyrogenicity is included, based on measuring the body temperature of rabbits after the introduction of test sterile liquids into the ear vein. Sampling is carried out in the same way as in the toxicity test. The general article (GF XI, issue 2, pp. 183--185) specifies the requirements for experimental animals and the procedure for their preparation for testing. The test solution is tested on three rabbits (not albino), whose body weight differs by no more than 0.5 kg. Body temperature is measured by inserting a thermometer into the rectum to a depth of 5--7 cm. The test liquids are considered non-pyrogenic if the sum of elevated temperatures in three rabbits is equal to or less than 1.4°C. If this amount exceeds 2.2°C, then water for injection or injection solution is considered pyrogenic. If the sum of the temperature rises in three rabbits is between 1.5 and 2.2°C, the test is repeated in an additional five rabbits. The test fluids are considered non-pyrogenic if the sum of the temperature rises in all eight rabbits does not exceed 3.7°C. In private FS, other temperature deviation limits may be specified. Rabbits that were in the experiment can be used for this purpose again no earlier than 3 days later, if the solution introduced by them was non-pyrogenic. If the injected solution turned out to be pyrogenic, then rabbits can be reused only after 2-3 weeks. In SP XI, in comparison with SP X, a test for the reactivity of rabbits used for the first time for testing has been introduced, and the section on the possibility of their use for repeated tests has been clarified.

The recommended SP XI biological method is specific, but does not quantify the content of pyrogenic substances. Its significant disadvantages include the complexity and duration of testing, the need to keep animals, care for them, the complexity of preparing for testing, the dependence of the results on the individual characteristics of each animal, etc. Therefore, attempts were made to develop other methods for determining pyrogenicity.

Along with the determination of pyrogenicity in rabbits, a microbiological method is used abroad, based on counting the total number of microorganisms in the studied dosage form before its sterilization. In our country, a simple and accessible method for the detection of pyrogens has been proposed, based on the selective identification of gram-negative microorganisms by the gel formation reaction using a 3% potassium hydroxide solution. The technique can be used at chemical and pharmaceutical enterprises.

An attempt was made to replace the biological method for determining pyrogenicity with a chemical one. Solutions containing pyrogens, after treatment with quinone, showed a negative reaction with tetrabromophenolphthalein. Pyrogenal with tryptophan in the presence of sulfuric acid forms a brown-raspberry color at a pyrogenal content of 1 μg or more.

The possibility of spectrophotometric determination of pyrogenic substances in the UV region of the spectrum was investigated. Solutions of the filtrate of pyrogen-containing cultures of microorganisms show a weak absorption maximum at 260 nm. In terms of sensitivity, the spectrophotometric method for determining pyrogens is 7-8 times inferior to the biological test on rabbits. However, if ultrafiltration is carried out before spectrophotometry, then due to the concentration of pyrogens, comparable results can be achieved by biological and spectrophotometric determinations.

After treatment with quinone, pyrogen solutions acquire a red color and a light absorption maximum appears at 390 nm. This made it possible to develop a photocolorimetric method for the determination of pyrogens.

The high sensitivity of the luminescent method created the prerequisites for its use for the determination of pyrogenic substances at concentrations up to 1*10 -11 g/ml. Methods have been developed for the luminescent detection of pyrogens in water for injection and in some injection solutions using the dyes rhodamine 6G and 1-anilino-naphthalene-8-sulfonate. The techniques are based on the ability of pyrogens to increase the intensity of the luminescence of these dyes. They allow you to get results comparable to the biological method.

The relative error of the spectrophotometric and luminescent determinations does not exceed ±3%. The chemiluminescent method is also used to determine the pyrogenicity of water for injection.

A promising method is polarography. It has been established that filtrates of pyrogenic cultures, even in a very dilute state, have a strong suppressive effect on the polarographic maximum of oxygen. On this basis, a method has been developed for the polarographic assessment of the quality of water for injection and some injection solutions.

Test for the content of histamine-like substances.

Parenteral medicinal products are subjected to this test. Perform it on cats of both sexes weighing at least 2 kg under urethane anesthesia. First, an anesthetized animal is injected with histamine, testing its sensitivity to this substance. Then, with an interval of 5 minutes, repeated injections (0.1 μg/kg) of the standard solution of histamine are continued until the same decrease in blood pressure is obtained with two successive injections, which is taken as standard. After that, with an interval of 5 minutes, the test solution is administered to the animal at the same rate as the histamine was administered. The drug is considered to have passed the test if the decrease in blood pressure after the introduction of the test dose does not exceed the response to the introduction of 0.1 µg/kg in the standard solution.

5 / 5 (votes: 1 )

Today, it is quite common to find low-quality medicines and dummy pills that cause the consumer to doubt their effectiveness. There are certain methods of drug analysis that allow to determine the composition of the drug, its characteristics with maximum accuracy, and this will reveal the degree of influence of the drug on the human body. If you have certain complaints about a drug, then its chemical analysis and objective opinion can be evidence in any legal proceeding.

What methods of drug analysis are used in laboratories?

To establish the qualitative and quantitative characteristics of a drug in specialized laboratories, the following methods are widely used:

Physical and physico-chemical, which help determine the melting and solidification temperature, density, composition and purity of impurities, find the content of heavy metals.
Chemical, determining the presence of volatile substances, water, nitrogen, the solubility of the medicinal substance, its acid, iodine number, etc.
Biological, allowing you to test the substance for sterility, microbial purity, the content of toxins.

Methods for the analysis of medicines will make it possible to establish the authenticity of the composition declared by the manufacturer and determine the slightest deviations from the norms and production technology. The laboratory of ANO "Center for Chemical Expertise" has all the necessary equipment for an accurate study of any type of medicine. Highly qualified specialists use a variety of methods for analyzing medicines and will provide an objective expert opinion in the shortest possible time.

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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.

Chapter 1. Basic Principles of Pharmaceutical Analysis

1.1 Pharmaceutical analysis criteria

At various stages of pharmaceutical analysis, depending on the tasks set, criteria such 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 can be detected by this method with a given confidence level. 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 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.

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 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 ,5)%; 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.

1.3 General principles for testing the identity of medicinal substances

1.4 Sources and causes of poor quality of medicinal substances

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, drying mode and temperature, which fluctuates 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 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".

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.).

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

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 assessed 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 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 medicinal product 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 block-heater, 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 (indicated 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 mm Hg. (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, vol. 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 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 determined.

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, monobasic 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. 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 standard 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 determined. 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. The identification of inorganic medicinal substances consists in the detection, by means of chemical reactions, of the cations and anions that make up the molecules. The 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 permissible limits for the content of impurities.

Chemical methods have proved to be the most reliable and effective, they make it possible 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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Physico-chemical or instrumental methods of analysis

Physico-chemical or instrumental methods of analysis are based on the measurement of the physical parameters of the analyzed system, which occur or change during the course of the analytical reaction, using instruments (instruments).

The rapid development of physical and chemical methods of analysis was due to the fact that the classical methods of chemical analysis (gravimetry, titrimetry) could no longer satisfy the numerous requests of the chemical, pharmaceutical, metallurgical, semiconductor, nuclear and other industries that required an increase in the sensitivity of methods up to 10-8 - 10-9%, their selectivity and rapidity, which would make it possible to control technological processes according to chemical analysis data, as well as to perform them automatically and remotely.

A number of modern physicochemical methods of analysis make it possible to simultaneously perform both qualitative and quantitative analysis of components in the same sample. The accuracy of the analysis of modern physicochemical methods is comparable to the accuracy of classical methods, and in some, for example, in coulometry, it is significantly higher.

The disadvantages of some physicochemical methods include the high cost of the instruments used, the need to use standards. Therefore, classical methods of analysis still have not lost their value and are used where there are no restrictions on the speed of analysis and where high accuracy is required at a high content of the analyzed component.

Classification of physical and chemical methods of analysis

The classification of physicochemical methods of analysis is based on the nature of the measured physical parameter of the analyzed system, the value of which is a function of the amount of substance. In accordance with this, all physicochemical methods are divided into three large groups:

Electrochemical;

Optical and spectral;

Chromatographic.

Electrochemical methods of analysis are based on the measurement of electrical parameters: current strength, voltage, equilibrium electrode potentials, electrical conductivity, amount of electricity, the values of which are proportional to the content of the substance in the analyzed object.

Optical and spectral methods of analysis are based on measuring parameters that characterize the effects of the interaction of electromagnetic radiation with substances: the intensity of the radiation of excited atoms, the absorption of monochromatic radiation, the refractive index of light, the angle of rotation of the plane of a polarized light beam, etc.

All these parameters are a function of the concentration of the substance in the analyzed object.

Chromatographic methods are methods for separating homogeneous multicomponent mixtures into individual components by sorption methods under dynamic conditions. Under these conditions, the components are distributed between two immiscible phases: mobile and stationary. The distribution of the components is based on the difference in their distribution coefficients between the mobile and stationary phases, which leads to different rates of transfer of these components from the stationary to the mobile phase. After separation, the quantitative content of each of the components can be determined by various methods of analysis: classical or instrumental.

Molecular absorption spectral analysis

Molecular absorption spectral analysis includes spectrophotometric and photocolorimetric types of analysis.

Spectrophotometric analysis is based on the determination of the absorption spectrum or the measurement of light absorption at a strictly defined wavelength, which corresponds to the maximum of the absorption curve of the substance under study.

Photocolorimetric analysis is based on a comparison of the color intensity of the investigated colored and standard colored solutions of a certain concentration.

Molecules of a substance have a certain internal energy E, the components of which are:

Energy of motion of electrons Еel located in the electrostatic field of atomic nuclei;

Vibration energy of atomic nuclei relative to each other E col;

Energy of rotation of the molecule E vr

and mathematically expressed as the sum of all the above energies:

Moreover, if a molecule of a substance absorbs radiation, then its initial energy E 0 increases by the amount of energy of the absorbed photon, that is:

It follows from the above equality that the shorter the wavelength l, the greater the frequency of oscillations and, therefore, the greater E, that is, the energy imparted to the molecule of the substance when interacting with electromagnetic radiation. Therefore, the nature of the interaction of ray energy with matter, depending on the wavelength of light l, will be different.

The totality of all frequencies (wavelengths) of electromagnetic radiation is called the electromagnetic spectrum. The wavelength interval is divided into areas: ultraviolet (UV) approximately 10-380 nm, visible 380-750 nm, infrared (IR) 750-100000 nm.

The energy imparted to a substance molecule by UV and visible radiation is sufficient to cause a change in the electronic state of the molecule.

The energy of infrared rays is less, so it is only sufficient to cause a change in the energy of vibrational and rotational transitions in a molecule of matter. Thus, in different parts of the spectrum it is possible to obtain different information about the state, properties and structure of substances.

Radiation Absorption Laws

Spectrophotometric methods of analysis are based on two main laws. The first of them is the Bouguer-Lambert law, the second law is Beer's law. The combined Bouguer-Lambert-Beer law has the following formulation:

The absorption of monochromatic light by a colored solution is directly proportional to the concentration of the light-absorbing substance and the thickness of the solution layer through which it passes.

The Bouguer-Lambert-Beer law is the basic law of light absorption and underlies most photometric methods of analysis. Mathematically, it is expressed by the equation:

the value log I/I 0 is called the optical density of the absorbing substance and is denoted by the letters D or A. Then the law can be written as follows:

The ratio of the intensity of the monochromatic radiation flux passing through the test object to the intensity of the initial radiation flux is called the transparency, or transmission, of the solution and is denoted by the letter T: T = I/I 0

This ratio can be expressed as a percentage. The value of T, which characterizes the transmission of a layer 1 cm thick, is called the transmission coefficient. Optical density D and transmission T are related by the relationship

D and T are the main quantities characterizing the absorption of a solution of a given substance with a certain concentration at a certain wavelength and thickness of the absorbing layer.

Dependence D(С) is rectilinear, and Т(С) or Т(l) is exponential. This is strictly observed only for monochromatic radiation fluxes.

The value of the extinction coefficient K depends on the method of expressing the concentration of the substance in the solution and the thickness of the absorbing layer. If the concentration is expressed in moles per liter, and the layer thickness is in centimeters, then it is called the molar extinction coefficient, denoted by the symbol e and is equal to the optical density of a solution with a concentration of 1 mol / l, placed in a cuvette with a layer thickness of 1 cm.

The value of the molar light absorption coefficient depends on:

From the nature of the solute;

Wavelengths of monochromatic light;

Temperatures;

The nature of the solvent.

Reasons for non-observance of the Bouger-Lambert-Beer law.

1. The law is derived and valid only for monochromatic light, therefore, insufficient monochromatization can cause a deviation of the law, and the more so, the less monochromatization of light.

2. Various processes can occur in solutions that change the concentration of an absorbing substance or its nature: hydrolysis, ionization, hydration, association, polymerization, complex formation, etc.

3. The light absorption of solutions significantly depends on the pH of the solution. When the pH of the solution changes, the following can change:

The degree of ionization of a weak electrolyte;

The form of existence of ions, which leads to a change in light absorption;

The composition of the resulting colored complex compounds.

Therefore, the law is valid for highly dilute solutions, and its scope is limited.

visual colorimetry

The color intensity of solutions can be measured by various methods. Among them, subjective (visual) methods of colorimetry and objective, that is, photocolorimetric, are distinguished.

Visual methods are such methods in which the assessment of the color intensity of the test solution is done with the naked eye. With objective methods of colorimetric determination, photocells are used instead of direct observation to measure the color intensity of the test solution. The determination in this case is carried out in special devices - photocolorimeters, so the method is called photocolorimetric.

Visible light colors:

Visual methods include:

- method of standard series;

- method of colorimetric titration, or duplication;

- equalization method.

Standard series method. When performing analysis using the standard series method, the color intensity of the analyzed colored solution is compared with the colors of a series of specially prepared standard solutions (at the same layer thickness).

Method of colorimetric titration (duplication) is based on comparing the color of the analyzed solution with the color of another solution - the control. The control solution contains all components of the test solution, with the exception of the analyte, and all the reagents used in the preparation of the sample. A standard solution of the analyte is added to it from the burette. When so much of this solution is added that the color intensities of the control and analyzed solutions become equal, it is considered that the analyzed solution contains the same amount of the analyte as it was introduced into the control solution.

Adjustment method differs from the visual colorimetric methods described above, in which the similarity of the colors of the standard and test solutions is achieved by changing their concentration. In the equalization method, the similarity of colors is achieved by changing the thickness of the layers of colored solutions. For this purpose, when determining the concentration of substances, drain and dip colorimeters are used.

Advantages of visual methods of colorimetric analysis:

The determination technique is simple, there is no need for complex expensive equipment;

The eye of the observer can evaluate not only the intensity, but also the shades of the color of the solutions.

Flaws:

It is necessary to prepare a standard solution or a series of standard solutions;

It is impossible to compare the color intensity of a solution in the presence of other colored substances;

With a long comparison of the color intensity of the human eye, it gets tired, and the error in the determination increases;

The human eye is not as sensitive to small changes in optical density as photovoltaic devices, so it is not possible to detect differences in concentration up to about five relative percent.

Photoelectrocolorimetric methods

Photoelectrocolorimetry is used to measure the absorption of light or the transmission of colored solutions. Instruments used for this purpose are called photoelectrocolorimeters (PEC).

Photoelectric methods for measuring color intensity involve the use of photocells. In contrast to devices in which color comparisons are made visually, in photoelectric colorimeters, the receiver of light energy is a device - a photocell. This device converts light energy into electrical energy. Photocells make it possible to carry out colorimetric determinations not only in the visible, but also in the UV and IR regions of the spectrum. The measurement of light fluxes using photoelectric photometers is more accurate and does not depend on the features of the observer's eye. The use of photocells makes it possible to automate the determination of the concentration of substances in the chemical control of technological processes. As a result, photoelectric colorimetry is much more widely used in the practice of factory laboratories than visual.

On fig. 1 shows the usual arrangement of nodes in instruments for measuring the transmission or absorption of solutions.

Fig.1 The main components of devices for measuring radiation absorption: 1 - radiation source; 2 - monochromator; 3 - cuvettes for solutions; 4 - converter; 5 - signal indicator.

Photocolorimeters, depending on the number of photocells used in measurements, are divided into two groups: single-beam (one-arm) - devices with one photocell and two-beam (two-arm) - with two photocells.

The measurement accuracy obtained with single-beam FECs is low. In factory and scientific laboratories, photovoltaic installations equipped with two photocells are most widely used. The design of these devices is based on the principle of equalizing the intensity of two light beams using a variable slit diaphragm, that is, the principle of optical compensation of two light fluxes by changing the aperture pupil opening.

The schematic diagram of the device is shown in fig. 2. The light from the incandescent lamp 1 is divided by mirrors 2 into two parallel beams. These light beams pass through light filters 3, cuvettes with solutions 4 and fall on photocells 6 and 6", which are connected to galvanometer 8 according to a differential circuit. Slit diaphragm 5 changes the intensity of the light flux incident on photocell 6. Photometric neutral wedge 7 serves to attenuate light flux incident on the photocell 6 ".

Fig.2. Scheme of a two-beam photoelectrocolorimeter

Determination of concentration in photoelectrocolorimetry

To determine the concentration of analytes in photoelectrocolorimetry, the following are used:

Method for comparing the optical densities of standard and test colored solutions;

Method for determining the average value of the molar coefficient of light absorption;

Calibration curve method;

additive method.

Method for comparing the optical densities of standard and test colored solutions

For determination, prepare a standard solution of the analyte of known concentration, which approaches the concentration of the test solution. Determine the optical density of this solution at a certain wavelength D this. Then determine the optical density of the test solution D X at the same wavelength and at the same layer thickness. By comparing the optical densities of the test and reference solutions, an unknown concentration of the analyte is found.

The comparison method is applicable for single analyzes and requires observance of the basic law of light absorption.

Graduated Graph Method. To determine the concentration of a substance by this method, a series of 5-8 standard solutions of various concentrations is prepared. When choosing the range of concentrations of standard solutions, the following provisions are used:

* it should cover the area of possible measurements of the concentration of the test solution;

* the optical density of the test solution should correspond approximately to the middle of the calibration curve;

* it is desirable that in this range of concentrations the basic law of light absorption is observed, that is, the dependence graph is straightforward;

* The value of optical density should be in the range of 0.14 ... 1.3.

Measure the optical density of standard solutions and build a graph of dependence D(C) . Having defined D X of the investigated solution, according to the calibration graph, find FROM X (Fig. 3).

This method makes it possible to determine the concentration of a substance even in cases where the basic law of light absorption is not respected. In this case, a large number of standard solutions are prepared, differing in concentration by no more than 10%.

Rice. 3. The dependence of the optical density of the solution on the concentration (calibration curve)

Additive method- this is a kind of comparison method based on comparing the optical density of the test solution and the same solution with the addition of a known amount of the analyte.

It is used to eliminate the interfering influence of foreign impurities, to determine small amounts of the analyte in the presence of large amounts of foreign substances. The method requires obligatory observance of the basic law of light absorption.

Spectrophotometry

This is a photometric analysis method in which the content of a substance is determined by its absorption of monochromatic light in the visible, UV and IR regions of the spectrum. In spectrophotometry, in contrast to photometry, monochromatization is provided not by light filters, but by monochromators, which make it possible to continuously change the wavelength. As monochromators, prisms or diffraction gratings are used, which provide a significantly higher monochromaticity of light than light filters, so the accuracy of spectrophotometric determinations is higher.

Spectrophotometric methods, in comparison with photocolorimetric methods, allow solving a wider range of problems:

* carry out quantitative determination of substances in a wide range of wavelengths (185-1100 nm);

* carry out quantitative analysis of multicomponent systems (simultaneous determination of several substances);

* determine the composition and stability constants of light-absorbing complex compounds;

* determine the photometric characteristics of light-absorbing compounds.

Unlike photometers, the monochromator in spectrophotometers is a prism or a diffraction grating, which allows you to continuously change the wavelength. There are instruments for measurements in the visible, UV and IR regions of the spectrum. Schematic diagram of the spectrophotometer is practically independent of the spectral region.

Spectrophotometers, like photometers, are single- and double-beam. In double-beam instruments, the light flux is somehow bifurcated either inside the monochromator or after exiting it: one stream then passes through the test solution, the other through the solvent.

Single-beam instruments are especially useful when performing quantitative determinations based on optical density measurements at a single wavelength. In this case, the simplicity of the device and the ease of operation represent a significant advantage. The high speed and convenience of measurements when working with two-beam instruments are useful in qualitative analysis, when optical density must be measured over a wide range of wavelengths to obtain a spectrum. In addition, a two-beam device can be easily adapted for automatic recording of a continuously changing optical density: in all modern recording spectrophotometers, it is a two-beam system that is used for this purpose.

Both single and double beam instruments are suitable for visible and UV measurements. Commercially available IR spectrophotometers are always based on a two-beam design, as they are typically used to sweep and record a large region of the spectrum.

Quantitative analysis of one-component systems is carried out by the same methods as in photoelectrocolorimetry:

The method of comparing the optical densities of the standard and test solutions;

Method of determination by the average value of the molar coefficient of light absorption;

By the calibration curve method,

and has no distinguishing features.

Spectrophotometry in Qualitative Analysis

Qualitative analysis in the ultraviolet part of the spectrum. Ultraviolet absorption spectra usually have two or three, sometimes five or more absorption bands. For unambiguous identification of the substance under study, its absorption spectrum in various solvents is recorded and the data obtained are compared with the corresponding spectra of similar substances of known composition. If the absorption spectra of the substance under study in different solvents coincide with the spectrum of a known substance, then it is possible with a high degree of probability to conclude that the chemical composition of these compounds is identical. To identify an unknown substance by its absorption spectrum, it is necessary to have a sufficient number of absorption spectra of organic and inorganic substances. There are atlases that list the absorption spectra of very many, mainly organic substances. The ultraviolet spectra of aromatic hydrocarbons have been especially well studied.

When identifying unknown compounds, attention should also be paid to the absorption intensity. Very many organic compounds have absorption bands, the maxima of which are located at the same wavelength l, but their intensity is different. For example, in the spectrum of phenol, there is an absorption band at n = 255 nm, for which the molar absorption coefficient at the absorption maximum e max= 1450. At the same wavelength, acetone has a band for which e max = 17.

Qualitative analysis in the visible part of the spectrum. The identification of a colored substance, such as a dye, can also be carried out by comparing its absorption spectrum in the visible part with the spectrum of a similar dye. The absorption spectra of most dyes are described in special atlases and manuals. From the absorption spectrum of the dye, one can draw a conclusion about the purity of the dye, because the spectrum of impurities has a number of absorption bands that are absent in the spectrum of the dye. From the absorption spectrum of a mixture of dyes, one can also draw a conclusion about the composition of the mixture, especially if the spectra of the components of the mixture contain absorption bands located in different regions of the spectrum.

Qualitative analysis in the infrared region of the spectrum

The absorption of IR radiation is associated with an increase in the vibrational and rotational energies of the covalent bond, if it leads to a change in the dipole moment of the molecule. This means that almost all molecules with covalent bonds are to some extent capable of absorbing in the IR region.

The infrared spectra of polyatomic covalent compounds are usually very complex: they consist of many narrow absorption bands and are very different from conventional UV and visible spectra. The differences stem from the nature of the interaction between the absorbing molecules and their environment. This interaction (in condensed phases) affects the electronic transitions in the chromophore, so the absorption lines broaden and tend to merge into broad absorption bands. In the IR spectrum, on the contrary, the frequency and absorption coefficient corresponding to a single bond usually change little with a change in the environment (including changes in other parts of the molecule). The lines also expand, but not enough to merge into a strip.

Usually, when plotting IR spectra, the transmission as a percentage is plotted along the y-axis, and not the optical density. With this method of plotting, the absorption bands look like troughs on the curve, and not like maxima on the UV spectra.

The formation of infrared spectra is associated with the vibrational energy of molecules. Vibrations can be directed along the valence bond between the atoms of the molecule, in which case they are called valence. There are symmetrical stretching vibrations, in which the atoms vibrate in the same directions, and asymmetric stretching vibrations, in which the atoms vibrate in opposite directions. If vibrations of atoms occur with a change in the angle between the bonds, they are called deformation vibrations. Such a division is very conditional, because during stretching vibrations, the deformation of the corners occurs to one degree or another, and vice versa. The energy of bending vibrations is usually less than the energy of stretching vibrations, and the absorption bands due to bending vibrations are located in the region of longer waves.

Vibrations of all atoms of a molecule cause absorption bands that are individual for the molecules of a given substance. But among these vibrations, vibrations of groups of atoms can be distinguished, which are weakly related to vibrations of atoms in the rest of the molecule. The absorption bands due to such vibrations are called characteristic bands. They are observed, as a rule, in the spectra of all molecules in which these groups of atoms are present. An example of characteristic bands are the bands at 2960 and 2870 cm -1 . The first band is due to asymmetric stretching vibrations of the C-H bond in the methyl group CH 3, and the second is due to symmetrical stretching vibrations of the C-H bond of the same group. Such bands with a small deviation (±10 cm -1) are observed in the spectra of all saturated hydrocarbons and in general in the spectrum of all molecules in which there are CH 3 groups.

Other functional groups can affect the position of the characteristic band, and the frequency difference can be up to ±100 cm -1 , but such cases are few and can be taken into account on the basis of literature data.

Qualitative analysis in the infrared region of the spectrum is carried out in two ways.

1. Remove the spectrum of an unknown substance in the region of 5000-500 cm -1 (2 - 20 microns) and look for a similar spectrum in special catalogs or tables. (or using computer databases)

2. In the spectrum of the substance under study, characteristic bands are sought, by which one can judge the composition of the substance.

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Chemical methods of drug research. Methods of drug analysis

Introduction

Chapter 1. Basic Principles of Pharmaceutical Analysis

1.1 Pharmaceutical analysis criteria

1.2 Errors in Pharmaceutical Analysis

1.3 General principles for testing the identity of medicinal substances

1.4 Sources and causes of poor quality of medicinal substances

1.5 General requirements for purity tests

What methods of drug analysis are used in laboratories?

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Introduction

Chapter 1. Basic Principles of Pharmaceutical Analysis

1.1 Pharmaceutical analysis criteria

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

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.

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.

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.

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.

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 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.

1.3 General principles for testing the identity of medicinal substances

1.4 Sources and causes of poor quality of medicinal substances

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

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".

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

1.5 General requirements for purity tests

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.

Chapter 2. Physical Methods of Analysis

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

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 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.

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

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

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.

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

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.

3.2 Gravimetric (weight) method

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

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