Technique
Thin-Layer Chromatography as an Analytical Tool


Qualitative Analysis


Quantitative Analysis





Thin-Layer Chromatography as a Preparative Tool

Thin-layer chromatography of lipids

O-linked oligosaccharides of mouse egg ZP3 account for its sperm receptor activity

Dünnschicht-Chromatographie von Aminosäurederivaten auf Kieselgel G. N-(2,4-Dinitrophenyl)-Aminosäuren und 3-Phenyl-2-thiohydantoine

— A 100,000 g supernatant fraction from rat brain that was passed through a column of Sephadex G-25-40 was able, after addition of some factors, to incorporate [I4C]arginine (apparent Km= 5 μM) and [14C]tyrosine (apparent Km= 20 μM) into its own proteins. The factors required for the incorporation of [14C]arginine were: ATP (optimal concentration = 0-25-2 μM) and Mg2+ (optimal concentration 5 mM). For the incorporation of [I4C]tyrosine the required factors were: ATP (apparent Km= 0-75 μM), Mg2+ (optimalconcentration 8-16 mM) and K+ (apparent Km= 16 mM). Addition of 19 amino acids did not enhance these incorporations. Optimal pHs were: for [14C]arginine and [14C]tyrosine, respectively, 7-4 and 7-0 in phosphate buffer and 7–9 and 7-3-8-1 in tris-HCl buffer. Pancreatic ribonuclease abolished the incorporation of [14C]arginine but had practically no effect in the incorporation of [14C]tyrosine. Furthermore, [14C]arginyl-tRNA was a more effective donor of arginyl groups than [14C]arginine, whereas [14C]tyrosyl-tRNA was considerably less effective than [14C]tyrosine. The incorporations of [14C]arginine and [14C]tyrosine into brain proteins were from 25- to 2000-fold higher than for any other amino acid tested (12 in total). In brain [14C]arginine incorporation was higher than in liver and thyroid but somewhat lower than in kidney. In comparison to brain, the incorporation of [14C]tyrosine was negligible in liver, thyroid or kidney. Kinetic studies showed that the macromolecular factor in the brain preparation was complex. The protein nature of the products was inferred from their insolubilities in hot TCA and from the action of pronase that rendered them soluble. [14C]Arginine was bound so that its a-amino group remained free. Maximal incorporation of [14C]tyrosine in brain of 30-day-old rats was about one-third of that in the 5-day-old rat. The changes with postnatal age in the incorporation of [14C]arginine were not statistically significant.

A soluble preparation from rat brain that incorporates into its own proteins [14c]arginine by a ribonuclease‐sensitive system and [14c]tyrosine by a ribonuclease‐insensitive system

The pharmacological properties of narcotic analgesics have made the identification of such compounds very important in drug control and in research in medicine, pharmacy, and criminology. In these fields the requirements of the analysis with respect to sensitivity and specificity vary greatly as do, consequently, the methods employed. A detailed description of all these methods is outside the scope of this review; therefore, brief mention and references will be given to those that are already well documented with fuller detail reserved for the more recently developed methods. The physicochemical methods for identification are obviously applicable to all compounds possessing the properties, or combination thereof, upon which the analysis is based. Therefore, evaluation of the specificity of the analysis for a given compound is of utmost importance.

Methods of Chemical Analysis

Amino acid separation with 6 different solvents on chromatoplates prepared from Kieselgel G (‘Merck’) is described. As demonstrated in Figur 8 and discussed in the text, this technique is equal or superior to paper chromatography.

M. Brenner

Papers

Dünnschicht-Chromatographie von Aminosäuren