Page 141 - phytochemistry general program

The chromatogram is placed in contact with a sheet of X-ray film, which after a
certain exposure time and development is found to have been blackened in the

areas corresponding to the active spots.

Evaluation/Analysis

(1) Visual estimation (Visual comparison of spots)
i. Test solution as well as standard solutions. (duplicates)
ii. After development and detection are completed, the size of the spot area is
evaluated, and the color intensity of the sample spot is compared with the dilution

series of standards.
(2) Physical measurement of colored spots
Densitometers of various types are specially made for the easy scanning of

chromatographic strips.
(3) Spot area measurements
The area of the spot is proportional to the concentration of the substance.
Needs round spots with a sharp boundary.
A calibration curve is used and usually of equal known volumes of both the sample

and standards are applied.
(4) Radioactive measurements
Using calibrated automatic scanning device. Autoradiography can be used.

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