The use of azo compounds in the spectrophotometric determination of drugs
1. Spectrophotometry
Spectrophotometry is a method of measuring how well a chemical absorbs light by measuring the intensity of light transmitted when a beam of light passes through a sample solution. The basic principle is that each compound absorbs or transmits light over a specific wavelength range. This measurement can also be used to measure the amount of a known chemical. Spectrophotometry is one of the most useful methods for quantitative analysis in various fields such as chemistry, physics, biochemistry, materials engineering and clinical applications. Any chemical compound that absorbs, transmits, or reflects light (electromagnetic radiation) over a specific wavelength range. In biochemistry, for example, it is used to identify enzyme-catalyzed reactions. In clinical applications, it is used to examine blood or tissue for clinical diagnosis. There are also many other techniques such as atomic absorption spectroscopy and atomic emission spectroscopy.
2. Spectrophotometer
A spectrophotometer is an instrument that measures the amount of photons (light intensity) absorbed after they have passed through a sample solution. With a spectrophotometer, a known chemical amount (concentrations) can also be determined by measuring the intensity of the detected light.
3. Chemical derivation
Since many compounds show either very weak or no absorption in the UV or visible regions, a number of methods using chemical derivatization have been developed. These methods usually involve the addition of an organic reagent, which forms a complex with a strong absorbance. The final stage of measurement is very similar to that of the direct methods. With this technique, proper reagent selection can greatly enhance sensitivity and selectivity. One of these derivatives are azo compounds.
4. azo compounds
Azo is an organic nitrogenous compound with an N=N- double bond. An azo compound has the formula (R-N = N-R-), where R- and R may be the same or different combinations. An azo group (-N = N-) is a bridging group that connects two moieties each on the bridge side.
4.1 Diazonium salts
HONO nitrous acid usually produces different kind of products by reaction with amines. Diazonium salts are formed when nitrous acid undergoes a reaction with primary aromatic amines. Aliphatic diazonium ions can also form when aliphatic primary amines react with nitrous acid, but they are unstable, even in solution. Aromatic diazonium salts are more stable than aliphatic diazonium salts.
4.2 Preparation of diazonium salts
Diazonium salts are prepared by treating nitrous acid (HON = O) with amine at about 0°C.
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Figure 1. Scheme of the azo reaction mechanism
4.3 Reactions of diazonium salts
Diazonium salts can react with other aromatic compounds to form intensely colored products called azo dyes. Azo dyes can be yellow, orange, red or green. Some azo species change color under acidic and basic conditions and can be used as indicators such as methyl orange.
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Figure 2. Scheme of the reaction mechanism by which diazonium salts react with
aromatic compounds
Diazonium salts are weak electrophiles, so they interact only with the aromatic system. Hydroxyl and amino groups are the most commonly electron releasing groups, therefore when they are present in an aromatic system, they would make a highly activated aromatic system for electrophilic attack. There are several methods in which drugs (which contain an amino group or a nitro group after being reduced to an amino group) are estimated using an azo-coupling reaction of the drug with an appropriate reagent. The following examples illustrate some of these drugs.
1. Husam S. Khalaf et al. have developed simple, sensitive, accurate and inexpensive spectrophotometric methods for the determination of sulfamethoxazole (SMZ) in pure dose form. This method is based on the conversion of the primary amine group of sulfamethoxazole using sodium nitrite and hydrochloric acid to dezanium salt followed by coupling with diphenylamine in acidic media to obtain a stable blue dye that showed an absorption maximum (λmax) at 530 nm. This method obeyed Beer's law at concentrations of 0.5-12.0 µg/mL.
2.Kasim M.H. Al-Abbasi proposed a spectrophotometric method for the determination of tetracycline in aqueous solution. This method is based on coupling of tetracycline with para-nitroaniline reagent in basic medium to form a soluble violet dye, which is stable and exhibits maximum absorption at 569 nm. The method obeys Beer's law over a concentration range of 2-400 µg/mL.
3. Luma T. Daood has developed a spectrophotometric method for the determination of amounts of dapsone. This method was based on react dapsone with phloroglucinol as a coupling reagent in basic medium, to form a stable, water-soluble yellow dye that showed maximum absorption at 435 nm. The method obeys Beer's law over a concentration range between 10-250 µg/25mL.
4. Khawla Salman Abd-Alrassol proposed a rapid, accurate and controlled spectrophotometric method for the determination of Nitrofurantoin. The method depends on the reduction of the drug by zinc powder in an acidic medium of hydrochloric acid followed coupling it with 8-hydroxyquinoline to give a stable colored product, soluble in water and has a maximum absorption at 365 nm. The method obeys Beer's law with a concentration range between 0.5-25 µg/ml
5. Ali I. Abdullah and Sumayha M. Abass proposed a simple and accurate method for the determination of furosemide based on the conversion of a secondary amine into a primary amine by acidolysis and then coupling with resorcino as a coupling agent in aqueous media at pH 13. The reaction gave a light orange color product It showed maximum absorption at 430 nm. The method obeyed Beer's law with a concentration between (0.25-2.5) µg/ml.
6. Khawla Salman Abd-Alrassol et al. proposed a rapid and sensitive spectrophotometric method for the determination of the Gabapentin drug in pharmaceutical preparations. The method is based on the coupling reaction of Gabapentin with 8-hydroxyquinoline to form an olive colored azo dye which gave maximum absorption at 365 nm.