In an experiment it is found that the optical density of a solution of compound X was 0.7 when measured at 400nm. in a 1cm glass cuvette. The solution had been prepared by dissolving 35mg of X in 50 ml of water. What is the molar extinction coefficient of X at 400nm? (mol. wt. of X is 450)
(20% of the marks)
(ii) Measurements of lactate dehydrogenase (LDH) activity were carried out on plasma from an adult patient admitted to hospital with a suspected heart attack. The assay was carried out in the direction:
Pyruvate + NADH lactate + NAD+
The oxidation of NADH was observed spectrophotometrically at 340nm in a 1cm cuvette.
Patient plasma samples were prepared by adding 0.3ml of plasma to 2ml of a suitable buffer and then adding a further 1ml of a solution to the diluted plasma to stabilise it.
Assay mixtures were prepared in a final volume of 3ml and contained 0.5ml of the stabilised diluted plasma, 0.15mM NADH and 1mM pyruvate. The reaction was carried out at 25oC and started by adding the pyruvate last. The following readings were obtained:
Time after pyruvate addition/min Absorbance at 340nm
Given that the extinction coefficient of NADH is 6.22 x 103 l.mol.-1 cm.-1at 340nm, calculate the activity of LDH in the patient plasma in terms of mol. pyruvate reduced. min.-1. (ml plasma) –1
(80% of the marks)
Im not too sure where to start with this Q..Im guessing that for the first part, I use A=ecl, where l is 1, A is 0.7 and I work out c to then find e??