Quantitative colorimetric method

Use has been made by Grimmett and Richards58,59 of a quantitative, colorimetric method for determining imidazole derivatives.25... 

The determination of phosphate concentration in a washing powder requires first the hydrolysis of the tripolyphosphate components into the phosphate ion or its protonated forms. Then a quantitative colorimetric method based upon the absorption of the yellow complex of phosphate and ammonium vanado-molybdate can be used. A series of standard solutions was prepared. The complex was then developed in 10 ml aliquots of these solutions by adding a 5.0 ml aliquot of an ammonium vanadomolybdate solution. 

Sandler, M. and Ruthven, C. R. J., Quantitative colorimetric method for estimation of 3-methoxy-4-hydroxymandelic acid in urine value in diagnosis of phaeochromocytoma, Lancet, 1959, p. 114. 

Almost half of the known types of polysaccharides contain uronic acid units. Uronic acid residues may be determined by decarboxylation with 12% hydrochloric acid and the evolved carbon dioxide determined by absorption (4) or by manometric measurement (S). A micro-quantitative colorimetric method employing carbazole has been developed for uronic acids (6), (See also Part II of this chapter and Chapter XI.)... 

Schwartz, D. P. (1985). Quantitative colorimetric method for sulfamethazine in swine feeds. J. Assoc. Off. Anal. Chem. 68 214-217. 

Cole similarly employs the pink colour produced between chloroform and pyridine in the presence of strong sodium hydroxide as a quantitative colorimetric method for the determination of small amounts of chloroform. 

Contaminant by-products depend upon process routes to the product, so maximum impurity specifications may vary, eg, for CHA produced by aniline hydrogenation versus that made by cyclohexanol amination. Capillary column chromatography has improved resolution and quantitation of contaminants beyond the more fliUy described packed column methods (61) used historically to define specification standards. Wet chemical titrimetry for water by Kad Eisher or amine number by acid titration have changed Httle except for thein automation. Colorimetric methods remain based on APHA standards. 

Analytical Techniques. Sorbic acid and potassium sorbate are assayed titrimetricaHy (51). The quantitative analysis of sorbic acid in food or beverages, which may require solvent extraction or steam distillation (52,53), employs various techniques. The two classical methods are both spectrophotometric (54—56). In the ultraviolet method, the prepared sample is acidified and the sorbic acid is measured at 250 260 nm. In the colorimetric method, the sorbic acid in the prepared sample is oxidized and then reacts with thiobarbituric acid the complex is measured at - 530 nm. Chromatographic techniques are also used for the analysis of sorbic acid. High pressure Hquid chromatography with ultraviolet detection is used to separate and quantify sorbic acid from other ultraviolet-absorbing species (57—59). Sorbic acid in food extracts is deterrnined by gas chromatography with flame ionization detection (60—62). 

Sucrose quantitation has also been performed by colorimetric methods. However, in recent years, automated enzymatic analyzers and instmmental methods (eg, ion chromatography and hplc) have become increasingly popular, as they provide greater sensitivity and accuracy. Near infrared (nir) spectroscopy is currendy under evaluation as a tool for sucrose quantitation in sugar mills and food processing operations. 

Colorimetric Methods. Numerous colorimetric methods exist for the quantitative determination of carbohydrates as a group (8). Among the most popular of these is the phenol—sulfuric acid method of Dubois (9), which rehes on the color formed when a carbohydrate reacts with phenol in the presence of hot sulfuric acid. The test is sensitive for virtually all classes of carbohydrates. Colorimetric methods are usually employed when a very small concentration of carbohydrate is present, and are often used in clinical situations. The Somogyi method, of which there are many variations, rehes on the reduction of cupric sulfate to cuprous oxide and is appHcable to reducing sugars. 

By mixing suitable indicators together changes in colour may be obtained over a considerable portion of the pH range. Such mixtures are usually called universal indicators . They are not suitable for quantitative titrations, but may be employed for the determination of the approximate pH of a solution by the colorimetric method. One such universal indicator is prepared by dissolving 0.1 g of phenolphthalein, 0.2 g of methyl red, 0.3 g of methyl yellow, 0.4 g of... 

Quantitative estimates are available for the amoiint of hordenine in malt roots as well as in dried malt (23) For gramine, only a quantitative estimate for the amount found in acrospires is available (2U). Because non-specific colorimetric methods were used to determine the amounts of these alkaloids in malt, the quantitative data can only be considered approximate. 

Although widely used, colorimetric methods for determining total ur-onic acid and hexosamine are now regarded as providing, at best, only semi-quantitative estimates of heparin.42 Colorimetric reactions in a strong acid, such as the carbazole reaction for the uronic acids48 and the Morgan-Elson reaction for the hexosamines,49 are not specific for hepa-... 

Rao et al. [45] determined primaquine phosphate, in pharmaceutical dosage forms, by using a colorimetric method. Powdered tablets containing the equivalent of 100 mg of primaquine phosphate were heated with 25 mL of water for 10 min, the solution was cooled and filtered, and 10 mL portion of the filtrate was diluted 10-fold with water. A 5-mL portion of this solution was mixed with 5 mL of pH 5 buffer solution, 1 mL of 0.08%. amidopyrine solution in aqueous 95% alcohol and 2 mL of aqueous 0.1% sodium periodate. After 10 min, 0.5 mL of aqueous sodium metabisulfite solution was added and the absorbance was measured at 580 nm. Beer s law was obeyed between 4 and 43 pg/mL of primaquine phosphate. Recoveries were quantitative. 

Total flavonoid content. Quantitative analysis of flavonoids depends on the objective of the study. Colorimetric estimation of total flavonoid content is measured by the aluminum chloride colorimetric assay (Jia and others 1999 Chang and others 2002). The total flavonoid content measured in this way is normally expressed in equivalent values of a standard flavonoid, often catechin or quercetin equivalents. Not all subgroups of flavonoids can be quantified by colorimetric methods however, total anthocyanin content is determined using the pH-differentiation method (Boyles and others 1993). 

The excipients present in pharmaceutical formulations can and often do interfere with quantitation of APIs, limiting the applications of direct UV-vis measurement for analyzing formulated products due to its lack of specificity. To minimize the interference of excipients, colorimetric methods based on chemical reactions have been used for rapid determination of drug substances in pharmaceutical formulations although their role in pharmacopoeias has been greatly reduced.117-122... 

Introduction of microbiological methods for the determination of amino acids made possible the estimation of the amount of both free and combined amino acids in urine. Dunn et al. (D4), Thompson and Kirby (Tl), Eckhard and Davidson (El), and Woodson et al. (W3) estimated the amount of amino acids liberated in the course of acid or, as in the case of tryptophan determination, alkaline hydrolysis. Microbiological and colorimetric methods used for the determination of certain amino acids present very little opportunity for evaluating the proper quantitative relations between free and combined amino acids, since under the applied condition both combined and free amino acids are equally involved in the reaction. In 1949 Albanese et al. (A3) applied such methods to the quantitative determination of free and combined amino acids in the nondiffusible fraction of urine, and subjected the procedures to broad criticism from just this point of view. 

The simple colorimetric method is based on that originally published by Murphy and Riley (1962). Phosphates are colorless or pale in solution, so the phosphorus in the test solution is quantitatively converted to a colored compound, firstly using an acidic molybdate solution (ammonium molybdate in nitric acid) ... 

Acid-base (neutralization) reactions are only one type of many that are applicable to titrimetric analysis. There are reactions that involve the formation of a precipitate. There are reactions that involve the transfer of electrons. There are reactions, among still others, that involve the formation of a complex ion. This latter type typically involves transition metals and is often used for the qualitative and quantitative colorimetric analysis (Chapters 8 and 9) of transition metal ions, since the complex ion that forms can be analyzed according to the depth of a color that it imparts to a solution. In this section, however, we are concerned with a titrimetric analysis method in which a complex ion-forming reaction is used. 

Some enzymes with improved single amino acid specificity are commercially available. An example is phenylalanine dehydrogenase (EC 1.4.1.1), derived from bacterial sources, which acts on phenylalanine with the simultaneous conversion of NAD to NADH. Quantitation of the phenylalanine is based on determining the amount of NADH produced using standard procedures. In the direct methods, the absorbance at 340 nm is measured, whereas in the colorimetric methods, the reaction is coupled to an electron acceptor... 

Being a phenolic compound, isoxsuprine has been determined by a diversity of colorimetric methods. A colorimetric method was described for its determination in dosage forms, based on treatment with the nitrating mixture composed of 1 1 sulfuric and nitric acids [13], This mixture is heated at 100°C for 20 minutes, followed by the addition of acetone and NaOH. The yellow color produced is quantitated on the basis of its absorbance at 384 nm. 

Tartaric Acid. Quantitative measures of total tartrate are useful in determining the amount of acid reduction required for high acid musts and in predicting the tartrate stability of finished wines. Three procedures may be used. Precipitation as calcium racemate is accurate (85), but the cost and unavailability of L-tartaric acid are prohibitive. Precipitation of tartaric acid as potassium bitartrate is the oldest procedure but is somewhat empirical because of the appreciable solubility of potassium bi-tartrate. Nevertheless, it is still an official AO AC method (3). The colorimetric metavanadate procedure is widely used (4, 6, 86, 87). Tanner and Sandoz (88) reported good correlation between their bitartrate procedure and Rebeleins rapid colorimetric method (87). Potentiometric titration in Me2CO after ion exchange was specific for tartaric acid (89). 

Colorimetric method of identification is described in this volume of Encyclopedia under Color Reactions and Color Reagents and also in Ref 6 Quantitative method of analysis of Comp B is given in Ref 2, which also lists the following US Military Spec requirements for Comp B ... 

The four colorimetric methods for the detection and quantitation presented in this unit have withstood the test of time. They are all well-characterized robust assays that consistently work well. The methods were introduced over the past 15 to 50 years. They collectively represent the state of the art for colorimetric detection and quantitation of total proteins in the microgram to milligram range. 

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