Bioanalytical considerations for compounds containing free sulfhydryl groups

Abstract

Thiol-containing compounds pose bioanalytical challenge in several dimensions due to extreme reactivity of the sulfhydryl group. The development of robust bioanalytical methodology for thiol groups should address the aspects of adequate stabilization in the biological matrix and selectivity considerations. In this context, availability of plethora of thiol reagents provides ample opportunity to achieve the above goals. However, several considerations need to be factored into the decision-making of a suitable scheme for the thiol drugs under investigation. This review provides some critical insights to many such considerations that may be vital for development and validation of methods for thiol-containing drugs.

Engineering an affinity tag for genetically encoded cyclic peptides

“…By incorporating a cysteine as the nucleophilic amino acid within the cyclic peptide sequence, the thiol side chain could be employed not only to effect splicing, but also as a universal chemoselective handle for covalent attachment of peptides to a solid support. While previous results from this lab indicated potential difficulties in using a cysteine for SICLOPPS chemistry (Scott et al, 2001), the ''analytical nightmare'' (Srinivas and Mamidi, 2003) of dealing with thiol-containing peptides could be alleviated by employing suitable reducing agents or oxidation inhibitors in all buffers. A combination of tris[2-carboxyethylphosphine] (TCEP) as a reductant (Kirley, 1989) and ethylenediaminetetraacetic acid (EDTA) as a transition metal scavenger (Srinivas and Mamidi, 2003) was expected to serve as an effective thiol group preservative, while being fully compatible with the maleimide immobilization strategy.…”

“…While previous results from this lab indicated potential difficulties in using a cysteine for SICLOPPS chemistry (Scott et al, 2001), the ''analytical nightmare'' (Srinivas and Mamidi, 2003) of dealing with thiol-containing peptides could be alleviated by employing suitable reducing agents or oxidation inhibitors in all buffers. A combination of tris[2-carboxyethylphosphine] (TCEP) as a reductant (Kirley, 1989) and ethylenediaminetetraacetic acid (EDTA) as a transition metal scavenger (Srinivas and Mamidi, 2003) was expected to serve as an effective thiol group preservative, while being fully compatible with the maleimide immobilization strategy. Chemoselective capturing of cysteinecontaining peptides on such surfaces should display small molecules for detection with an enzyme conjugate, not unlike in a traditional immunosorbent format (i.e., ELISA).…”

LC‐MS in Drug Metabolism and Pharmacokinetics: A Pharmaceutical Industry Perspective

“…Isomerization can be prevented by controlling exposure to UV wavelengths of light, and by temperature control. Sulfhydryl compounds are often stabilized by reacting with a derivatizing agent to prevent its reaction with another − SH group [107] . In addition to chemical reactions, there are a variety of enzymes in biological matrix that can catalyze degradation of drugs and metabolites.…”

Bromide–sulfur interchange: Ion chromatographic determination of total reduced thiol levels in plasma

“…The latter would act as an indirect measure of total PSH and, as IC is a widely established technique within most bioanalytical environments, it should allow rapid technology transfer from research study to routine application. Numerous strategies have been employed for the detection of thiols and the various merits and limitations have been critically reviewed [12][13][14]. A core aim of method proposed within Scheme 1 is that it will provide greater simplicity than many of the assay systems previously reported.…”