Assays molar absorption coefficients

The molar absorption coefficient of the coloured product can be used in the quantitation of individual amino acids but this value varies from one amino acid to another and must be determined under the conditions of the assay. An accepted value, however, must be used in the quantitation of the total amino acids in a mixture when absorbance readings are normally taken at 570 nm. 

Standard curve A series of dilutions of bovine serum albumin in water are prepared containing between 0 and 100 pig protein in 1 ml volume and the assay is performed as described above on each dilution. The exact concentration of the bovine serum albumin stock solution is determined using the molar absorption coefficient of 45 000 for bovine serum albumin at 279 nm after diluting to approximately 0.2 mg/ml. 

Yeast alcohol dehydrogenase (ADH) has its highest activity at 30°, and this decreases sharply with increase of temperature above 50°. No activity is observed at 70°. This inactivation process can be monitored by measuring the remaining activity of the enzyme during the incubation at elevated temperatures. ADH from yeast is assayed by monitoring ethanol-dependent NAD reduction at 340 nm ADH activity is expressed as micromoles of NADH produced per minute with a molar absorption coefficient of 6.22 mA/ cm . The influence of molecular chaperonin on ADH inactivation can be examined as follows. 

The coenzyme activity of Alg-NADP was determined enzymatically by the glutathione reductase system (38). The assay mixture consisted of 250 pmol Tris-HCl buffer (pH 8.0), 2.7 [xmol EDTA, 13.2 pmol GSSG, an appropriate amount of Alg-NADP", and 20 units of GR in a total volume of 2.87 ml. The reaction was started by the addition of the enzyme and the reaction mixture was incubated at 30°C. Absorbance at 340 nm was measured with a double beam spectrophotometer. The reference contained all components except for Alg-NADP. In the calculation of the amount of Alg-NADPH produced, a molar absorption coefficient for NADPH of 6.22 X 10 l-mol -cm was used. 

Table 1. Colorimetric analysis of the sialic acids. Molar absorption coefficients The molar absorption coefficients are noted for the different sialic acids using the commonly used colorimetric assays. The chromophore was measured at the absorption maximum, noted at the head of each column. Abbreviations of the sialic acids are those used in chapter B, Table 1... 

BChl 663 with absorption maxima at 663 and 4-33 nm, like BChl c, was found recently in Prosthecochloris aestuarii by means of reversed-phase HPLC [8], and the presence of this pigment in other species of green sulfur bacteria has not been reported. BChl 663 was assayed at an amount of 10-15 molecules per reaction center and was supposed to be a likely candidate for the primary electron acceptor [8]. In our present study the BChl a/BChl 663 molar ratio is 9 5-11 9 (four independent measurements), where the extinction coefficient of BChl 663 is assumed to be equal to that of BChl c. Although we have not... 

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