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Indian Journal of Clinical Biochemistry logoLink to Indian Journal of Clinical Biochemistry
. 2008 Jun 11;23(2):117–122. doi: 10.1007/s12291-008-0028-0

Effect of pH on spectral characteristics of P5C-ninhydrin derivative: Application in the assay of ornithine amino transferase activity from tissue lysate

H Ravikumar 1, K S Devaraju 1,2, K Taranath Shetty 1,✉
PMCID: PMC3453081  PMID: 23105736

Abstract

Currently available method(s) for assaying pyrroline-5-carboxylate (P5C), an important intermediate metabolite of ornithine, proline and glutamate metabolic pathways, are cumbersome or not sensitive enough for microanalysis. The present study involving the synthesis of P5C followed by purity check, molecular mass (amu =113.1) determination by mass spectrometry and spectral characterization of P5C-ninhydrin derivative (λ max: 510 nm) confirmed the authenticity of the preparation. Studies on the effect of pH on spectral characteristics of P5C ninhydrin derivative demonstrated a significant change with respect to λ max (620 nm) and several ∼ 12 fold increase in molar extinction coefficient (ε: 1.96 × 105) in alkaline conditions (pH:7.0–8.0) as compared to the reported Molar ε of 1.65 × 104 at max λ 510 nm in ethanolic solution. The modified method, with the improved sensitivity, is adopted for the assay of ornithine amino transferase activity in WBC’s/platelets lysate(s) from human blood.

Key Words: Pyrroline-5-Carboxylate, Mass spectrometry, WBC’s/platelets lysate(s), Ornithine amino-transferase, Pyrroline-5-Carboxylate-Ninhydrin complex

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