4) Standard solutions with known concentration of bovine serum albumin (BSA) are

Question

4) Standard solutions with known concentration of bovine serum albumin (BSA) are prepared for a calibration curve to determine the unknown concentration of BSA in a solution. In Table 2 are reported the absorbance value measured for eight standard solutions at different BSA concentration values.

a) Identify the concentration range under which the Beer-Lambert law is applicable and determine the BSA concentration for an absorbance equal to 0.234.

b) Calculate the extinction coefficient for BSA.

Table 2. The absorbance values of BSA for different concentration values in an aqueous solution Unit Concentration [mg mL j 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 Absorbance [J 0.108 0.178 0.241 0.312 0.395 0.423 0.435 0.44

Explanation / Answer

According to Beer-Lambert Law

    A = bc

or, = A/cb

where, c=concentration in mol/lt

b=path length of absorbing solution in cm-1

A= absorbance or optical density

= molar extinction coefficient

when, c=0.2 mg /ml , A=0.108

so,    = 0.108/ 0.2 x b

Let us assume that, C= C1 when A = 0.234

C1 = 0.234/    x b   = (0.234 x 0.2 x b) / (0.108 x b) = 0.433 mg/ml (Answer of question a)

Now,   = 0.108/ 0.2 x b = 0.54 ml mg-1 cm-1 (Answer of question b) whbn cell length(b) = 1cm

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