The students conducted the assay for LDH activity using the two serum samples. Each cuvette (path length 1 cm) contained 3ml of a suitable assay buffer (including pyruvate as substrate). 20 microlitres of either serum was added to the cuvette and the absorbance values immediately recorded at the optimum wavelength for a period of 5 minutes (absorbance readings taken every 30 seconds).   Protein concentration of serum sample (mg/ml) Change in absorbance at optimum wavelength per minute Control serum (C) 8 -0.04 Diseased serum (D) 7.8 -0.6 1c. Using the molar absorption coefficient of NADH as 6220 M-1 cm-1, and by application of the Beer-Lambert law, estimate the enzyme activity in the two samples (C and D). Express activity as moles per second.  1d. Estimate the specific activity of the two samples (moles per second per microgram).

Fundamentals Of Analytical Chemistry
9th Edition
ISBN:9781285640686
Author:Skoog
Publisher:Skoog
Chapter27: Molecular Fluorescence Spectroscopy
Section: Chapter Questions
Problem 27.13QAP
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The students conducted the assay for LDH activity using the two serum samples. Each cuvette (path length 1 cm) contained 3ml of a suitable assay buffer (including pyruvate as substrate). 20 microlitres of either serum was added to the cuvette and the absorbance values immediately recorded at the optimum wavelength for a period of 5 minutes (absorbance readings taken every 30 seconds).

 

Protein concentration of serum sample

(mg/ml)

Change in absorbance at optimum wavelength per minute

Control serum (C)

8

-0.04

Diseased serum (D)

7.8

-0.6

1c. Using the molar absorption coefficient of NADH as 6220 M-1 cm-1, and by application of the Beer-Lambert law, estimate the enzyme activity in the two samples (C and D). Express activity as moles per second. 

1d. Estimate the specific activity of the two samples (moles per second per microgram).

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