Young test

In the wide field of applications, a visibility level VL = 3 - 60 is recommended. For our recognition task, we are obliged to take into account that our random conditions are far from the experimental conditions of the basic researches (Young test person with a high visus under ideal environmental conditions) [4]. Furthermore in our case we have a more difficult visual searching task. Parameter variations as the increase of presentation time from 0,2 to 1.0 s. and the detection propability from 50% to 100% are taken into account [5] In spite of the gliding variations of the parameters as well as the visibility level, for simplification let us assume VL = 10 as minimum requirement. 

The classical methods for detection and quantitation of racemization require analysis of the chiral purity of the product of a peptide-bond-forming reaction. For example, the Anderson test is used to explore a variety of reaction conditions for the coupling of Z-Gly-Phe-OH to H-Gly-OEt (Scheme 6). 9 The two possible enantiomeric tripeptides are separable by fractional crystallization, so that gravimetric analysis furnishes the racemization data. This procedure has a detection limit of 1-2% of the epimerized tripeptide. A modification by Kemp,1"1 utilizing 14C-labeled carboxy components, extends the detection limit by two to three orders of magnitude by an isotopic dilution procedure. The Young test 11 addresses the coupling of Bz-Leu-OH to H-Gly-OEt, and the extent of epimerization is determined by measurement of the specific rotation of the dipeptide product. 

Peptide synthesis. The anhydride effects peptide synthesis as shown in equation (I). Free acid groups arc buffered with a tertiary amine such as N-elhylmorpholine. The reaction is complete after 24 hours at room temperature. No raccmization is detected in the Anderson-Young test. The resulting alkylphosphonic acid derivative is soluble in water and easily removed. 

Peptide synthesis, The condensation of the tetrabutylammonium salt of N-benzyl-L-leudne (2) with glycine ethyl ester (3), the components in the Young test, in DMF in the presence of 1 as coupling reagent proceeds in 75-80% yield without racemization. 

The reagent promotes the coupling in high yields of acylamino acids with amino acid esters in benzene, ethanol, or THF at room temperature. No racemization was detected in the supersensitive Young test, and Bz-Leu-Gly-OEt, an 33.5°, was synthesized in 95% yield. Activation of the carboxyl group involves the transient formation of a mixed carbonic anhydride.3... 

In tests devised to determine the amount of racemization to be expected in peptide syntheses in which the carboxyl component is a di-or higher peptide, this method ranks below the racemization-resistant azide procedure but above almost all other standard methods. Using a very sensitive and accurate isotope dilution assay, 1 % racemization was observed in the formation of the Anderson test peptide (Z-Gly-Phe-Gly-OEt) and 7% racemization was observed in the formation of the Young test peptide (Bz-Lou-Gly-OEt) under optimized conditions. (The Young test was designed to exaggerate racemization problems, thus permitting more accurate studies of the effects of reaction condition variations.)... 

A similar reaction, but with an amine, results in amides. The yields in this reaction are usually high, and (he Young test indicates that racemization is slight in the case of peptides. DMF is a suitable solvent. ... 

Therefore the Young-test might lead to somewhat exaggerated estimates of racemization. This distortion is counterbalanced by the relative resistance of leucine to racemization, but an additional problem is created by the necessity to isolate the crude benzoyl-leucyl-glycine ethyl ester in excellent yield. If less than near quantitative yield is achieved in coupling or in recovery of the product then it remains possible that the unaccounted portion contains a not insignificant amount of the D-isomer. 

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