Assay British Pharmacopoeia

The British Pharmacopoeia specifies a biological assay for the sodium salt of rifamycia SV [14897-39-3]. It also specifies a spectrophotometric assay for rifampicia (201). The United States Pharmacopeia requires an hplc assay for rifampin (202). 

A method for the estimation of the total alkaloids and of non-phenolic alkaloids (emetine fraction) is given in the British Pharmacopoeia, 1932, Addendum VI, which requires the drug to contain not less than 2 per cent, of alkaloids, of which at least 55 per cent, must be non-phenolic bases, calculated as emetine. The British Pharmacopoeia also gives an assay process for emetine in emetine bismuth iodide, the form in which the drug is chiefly used in medicine it is required to contain not less than 25 and not more than 28 per cent, of emetine. In the United States Pharmacopoeia, XIII, both Cephcelis Ipecacuanha and C. acuminata are recognised and must contain not less than 2 per cent, of ether-soluble alkaloids. 

The British Pharmacopoeia has adopted the assay of chlorpromazine in acetone, using a non-aqueous titration with perchloric acid and methyl orange as the indicator [4], According to the described procedure, each mL of 0.1 M perchloric acid is equivalent to 0.03189 g of drug substance. For chlorpromazine hydrochloride, the BP has described its titration with NaOH in a 1 10 solution of 0.01 M HCl / ethanol. Determining the end point potentiometrically, each mL of 0.1 M sodium hydroxide is equivalent to 0.03558 g of substance. 

The 1980, 1988 and 1993 editions of the British Pharmacopoeia have adopted the triphenyltetrazolium chloride method, but with certain modifications as suggested by Johnson et al. [85], The method described in the following section is that recommended by the B.P. of 1988 [3], It should be noted that all solutions must be protected from light throughout the assay. 

The molar absorptivity (e) of a known molecule is constant under identical conditions of solvent, concentration and path length, and can be used to quantify the amount of a particular pharmaceutical in a tablet. Such assays form the basis of many quality assurance procedures in the pharmaceutical industry, and have been extensively used by the British Pharmacopoeia (B.P.). More recently, however, high-performance liquid chromatography (HPLC) has replaced UV analysis in many B.P. assays, as most industrial analyses routinely use HPLC. 

According to the British Pharmacopoeia 2002, the European and Indian Pharmacopoeias, and the Pharmacopoeia of the People s Republic of China, the assay for mefenamic acid is performed by a titration method. About 0.200 g of the substance to be assayed is dissolved with the aid of ultrasound in 100 mL of warm ethanol that has been previously neutralized to the phenol red endpoint. About 0.1 mL of phenol red solution is added and titrated with 0.1 M sodium hydroxide. Each milliliter of 0.1 M NaOH is equivalent to 24.13 mg of mefenamic acid [2, 5-7],... 

In some assays it is necessary to specify the minimum desirable resolution, since changes in the spectral bandwidth (or monochromator slit-width) can seriously affect the observed absorbance of sharp peaks. The British Pharmacopoeia (1980) requires that the spectral bandwidth employed should be such that further reduction does not lead to an increase in measured absorbance. This is particularly important for drugs that have aromatic or strongly-conjugated systems, e.g. diphenhydramine,... 

An interesting example is die assay of clonidine hydrochloride in injections and tablets (British Pharmacopoeia, 1980). In 0.01 Mhydrochloric acid, clonidine exhibits two sharp maxima ne - 272 nm and 279 nm, which are not suitable for precise measurement. However, clonidine forms an ion pair with bromothymol blue, and this can be readily extracted into chloroform for subsequent measurement of the broad maximum near 420 nm. Because of the intiinsic variabihty of reagents used in such methods, a pharmacopoeial reference standard is employed for cahbration. A similar policy is adopted for assays involving chemical modification of the drug, as in the tetrazolium assay for corticosteroids, the assay for folic acid involving hydrolysis, diazotisation, and coupling with N-(l-naphthyl)ethylenediamine, and the reaction of penicillins with imidazole and mercuric salts. 

The European and British Pharmacopoeias [2] use the mercurimetric titration assay described in Section 6.2.2 for assay of both the trihydrate and sodium salt. The method is also specified for determination of degradation products in the sodium salt monograph. 

Degradation of amoxicillin at pH 5.2 in the presence of cupric ion gives a product absorbing at 320nm. Measurement of this was used as the assay method in the 1973 British Pharmacopoeia [75], The product was... 

Before we consider topics such as the design of an assay, calculation of drug purity, and so on, it is useful to revise the units and terms chemists use for amount of substance and concentration. The fundamental unit of quantity or amount of substance used in chemistry is the mole. The mole is the amount of a substance (either elements or compounds) that contains the same number of atoms or molecules as there are in 12.0000 g of carbon-12. This number is known as the Avogadro number (after Amedeo Avogadro, an Italian chemist) or Avogadro s constant, and has the value 6.02 X 1023. When this amount of substance is dissolved in solvent (usually water) and made up to 1 litre, a 1 molar (1 m) solution is produced. In a similar way, if one mole of substance were made up to 2 litres of solvent, a 0.5 m solution would result, and so on. The litre is not the SI unit of volume but, along with the millilitre (mL), is still used in the British Pharmacopoeia. 

A duplicate determination is now carried out and the answer is compared to 100.2% w/w. Agreement is usually considered to be not more than 0.5% error between duplicates. Once duplicate determinations have been carried out, and agreement is obtained, the answers may be averaged and the British Pharmacopoeia consulted to see whether the sample complies. Not every sample assayed will comply there may be impurities present if, for example, the sample was old or had been adulterated. However, an analyst who has obtained duplicate results, in good agreement, should be confident to state that the sample does not comply with the BP limits. 

In the UK and Europe the Beer-Lambert equation tends to be used in what is called the absolute method of assay. In this procedure the absorbance is measured experimentally and the Beer-Lambert equation is solved for c, the drug concentration. For this reason, the British Pharmacopoeia and European Pharmacopoeia quote A values in drug monographs. 

The BP Commission can also call on expertise available in the British Pharmacopoeia laboratories situated in the premises of the Laboratory of the Government Chemist in West London. The BP laboratory carries out and validates assay procedures for the Commission and in addition, is responsible for the procurement, establishment, maintenance and sale of British Pharmacopoeia Chemical Reference Substances (BPCRS). These reference substances, as their name suggests, are authentic samples of a drug or decomposition product which are used as standards in a drug assay. The BP laboratory also fulfils an important forensic role in the control of counterfeit medicines. With the advent of the internet, the public can easily gain access to supplies of prescription-only medicines online. These medicines are often adulterated, contaminated or simply counterfeit, and comparison with authentic samples is necessary to ensure that the correct preparation is supplied. 

Also the British Pharmacopoeia 1980 describes an assay for atropine in atropine sulphate eye drops and tablets, as well as in Hyoscyamus dry extract. However, atropine was silanized with N,0-bis(trimethylsilyl)acetamide and trimethyldichiorosilane (4 1) and gas chromatographed on a 1.5 m long by 4 mm I.D. packed with 3 % 0V-17 bn "diatomaceous support" AWS at... 

Microbiological assay is a technique in which the potency or concentration of a compound is assessed by determining its effect on micro-organisms. The principles are discussed by Roberts and Boyce. Microbiological assay is a legal QC requirement for the assay of a number of antibiotics, in both the British Pharmacopoeia (BP) and United States Pharmacopoeia (USP). 

More than 220 producers of CRMs throughout the world produce today 12,000 20,000 materials with dif ferent matrixes, analytes and properties [4]. However, many testing (analytical) laboratories cannot find suitable CRMs in the market and develop in-house reference materials (IHRMs) themselves. Often IHRMs are developed in a laboratory to conserve the corresponding expensive CRMs. For example, a pharmaceutical company Chemagis Ltd. produces 30 active pharmaceutical ingredients steroids, benzodiazepines, antihistamines, hipolipidaemics, blood flow reactants, etc. Only for a few of them Mo-metasone Furoate, Fluticasone Propionate and Dobutamine Hydrochloride are of fi-cial reference standards for assay supplied by US, British and European Pharmacopoeias with prices of about 180 per unit (50 200 mg). Thus, to support its customers Chemagis is forced to develop IHRMs for assay as well as for impurities and related substances of each produced compound. Therefore, certification of such IHRMs that leads to traceable values is very important. 

British Pharmacepoeia (BP) and European Pharmacopoeia (EP) Applications of GC for the Assay Chromatographic Purity Identification Presence of Volatile Matter, Intermediates and Related Substances Organic Volatile Impurities Determination of Water Presence of Isomers and Racemate Ratios Determination of Alcohol and Miscellaneou s Uses of GC in Pharmaceutical Raw Materials and Dosage Forms. 

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