Page 112 - Color Atlas of Biochemistry

Enzymes 103

A. Principle of spectrophotometry

Monochroma- Monochroma-
White light tic light, tic light,
intensity 0 intensity

d

Absorption

Light source Monochromator Detector Instrument
Light Sample solution,
absorption concentration c

. .
Absorption A = –log I = ε c d Beer Lambert
I 0 law

B. Assay of lactate dehydrogenase activity

∆A ∆A ∆c
2.0 ε = ∆c ; ∆t · ε = ∆t = v ; v ≈ Activity
Lactate
Pyruvate 0.1 mM
NAD each
1.5 NADH
Absorption Addition 3 nkat

1.0
NADH of LDH 2 nkat

0.5 Absorption (340 nm) Lactate; NAD 1 nkat
Lactate
NAD Pyruvate ∆A
0
0
1. 240 280 320 360 400 2. ∆t Time
Wavelength (nm)

C. Enzymatic determination of glucose

O 2
Glucose-containing
sample solution
Glucose Glucono-
Glucose oxidase lactone
1.1.3.4 [FAD]
Absorption (440 nm) Enzymes

2 H O H O Colorless ∆A
2 2
2
precursor
∆A
∞
ε
∞
Green Peroxidase Colorless [Green dye] = — 0
[Green dye] = [Glucose]
dye 1.11.1.7 precursor
[Heme]
0 10 20
Time (min)
1. Reaction 2. Procedure

Koolman, Color Atlas of Biochemistry, 2nd edition © 2005 Thieme
All rights reserved. Usage subject to terms and conditions of license.