Stoichiometry of interaction between soluble (—)-dopa melanin and enantiomers of ephedrine by NMR spectroscopy

Synthetic soluble (—)-dopa melanin was prepared in deuteriated buffer, pH 8, by autooxidation of the precursor. At 6 mM of the precursor, the incorporation was over 90%. The changes in the line width measurements of N-CH₃ protons of enantiomers of ephedrine in the soluble melanin were quantified by NMR spectroscopy. The dissociation constants of (—)-1R,2S-ephedrine, (+)-1S,2R-ephedrine, (—)-1R,2R-ψ-ephedrine, and (+)-1S,2S-ψ-ephedrine were 11.7, 4.20, 3.60, and 4.80 mM, respectively. Since the concentration of (—)-dopa was known and since the conversion of (—)-dopa to indole units of melanin was considered as 1:1, the stoichiometry of the interaction between the drug and the indole unit was calculated. Based on the dissociation constants of the enantiomers, it appears that up to four molecules of (—)-ephedrine can interact with one indole unit of the melanin, while such a ratio for other isomers appear to be 2:1. The preference by indole units of melanin is stereoselective. © 1992 Wiley-Liss, Inc.     

Formation of glycine conjugate and (-)-(R)-enantiomer from (+)-(S)-2-phenylpropionic acid suggesting the formation of the CoA thioester intermediate of (+)-(S)-enantiomer in dogs.

It has been proposed that the chiral inversion of the 2-arylpropionic acids is due to the stereospecific formation of the (-)-R-profenyl-CoA thioesters which are putative intermediates in the inversion. Accordingly, amino acid conjugation, for which the CoA thioesters are obligate intermediates, should be restricted to those optical forms which give rise to the (-)-R-profenyl-CoA, i.e., the racemates and the (-)-(R)-isomers. We have examined this problem in dogs with respect to 2-phenylpropionic acid(2-PPA). Regardless of the optical configuration of 2-phenylpropionic acid administered, the glycine conjugate was the major urinary metabolite and this was shown to be exclusively the (+)-(S)-enantiomer by chiral HPLC. Both (-)-(R)- and (+)-(S)-2-phenylpropionic acid were present in plasma after the administration of either antipode, and further evidence of the chiral inversion of both enantiomers was provided by the presence of some 25% of the opposite enantiomer in the free 2-phenylpropionic acid and its glucuronide excreted in urine after administration of (-)-(R)- and (+)-(S)-2-phenylpropionic acid. The (+)-(S)-enantiomer underwent chiral inversion to the (-)-(R)-antipode when incubated with dog hepatocytes. These data suggests that both enantiomers of 2-phenylpropionic acid are substrates for canine hepatic acyl CoA ligase(s) and thus undergo chiral inversion, but that the CoA thioester of only (+)-(S)-2-phenylpropionic acid is a substrate for the glycine N-acyl transferase. These studies are presently being extended to the structure and species specificity of the reverse inversion and amino acid conjugation of profen NSAIDs.     

The Effects of Surfactants on Lipase-Catalysed Hydrolysis of Esters: Activities and Stereoselectivity

The effect of surfactants on the hydrolysis of prochiral and chiral substrates by crude and purified porcine pancreatic lipase (PPL, EC 3.1.1.3)) has been studied. Rather than accelerating the reactions, surfactants slowed down (“inhibited”) the reactions relative to the rate in the absence of surfactant. Surfactants varied in the extent to which the reaction was inhibited. With the crude enzyme there was a correlation between degree of inhibition and the optical purity of the product of hydrolysis of an achiral diester substrate 1. There was no special effect associated with use of surfactants in the concentration range corresponding to critical micelle formation, nor was there any increase in rate of reaction when stable emulsions were formed by using mixtures of surfactants to generate an appropriate hydrophile-lipophile (HLB) balance. A study of the effect of sodium dodecyl sulphate (SDS) on the hydrolysis of the diester 1 by crude PPL showed that the rate of the reaction steadily decreased with increasing surfactant concentration, but that the optical purity of the product first fell and then rose gain, an effect attributed to the differential denaturing action of the surfactant on at least three hydrolytic enzymes. In general, there would seem to be no advantage to be gained from the use of surfactants in the hydrolysis by PPL of compounds of low water solubility; the use of an immiscible co-solvent is more effective.     

Stereoselectivity in β-cyclodextrin complexation of 1,4-dihydropyridine derivatives

Structure–interaction relationships, stereoselectivity, and solubility enhancement in inclusion compexation of β-cyclodextrins (CDs) with some racemic and enantiomerically pure 1,4-dihydropyridine derivatives (DHPs) were investigated. 1:1 and 1:2 (mole ratio) complexes were prepared and characterized by X-ray powder diffraction, differential scanning calorimetry (DSC), MS-FAB spectrometry, ¹H-NMR spectroscopy, water and phase solubility. The solubility studies have revealed different complexation equilibria for optically pure DHP enantiomers, and corresponding racemic mixtures in water solutions. By means of ¹H-NMR chemical shift measurements, the inclusion of aromatic fragments of racemic and enantiomerically pure DHP molecules within the cavities of different CDs was elucidated. Considerable stereoselectivity in complexation interactions was observed. The results indicate the potential use of cyclodextrins as chiral selectors for enantiomeric resolution of 1,4-DHP calcium antagonists. © 1993 Wiley-Liss, Inc.     

Chiral symmetry breaking: Experimental results and computer analysis of a liquid-phase autoxidation

The autoxidation of tetralin is treated as a model reaction system to define the applicability of stereospecific autocatalysis. This concept, predicting a spontaneous amplification of enantiomeric excess generated by an autocatalytic chemical reaction, is used in several theoretical models as an explanation for the origin of natural optical activity. The reaction system investigated obeys the basic criteria of these models: a chiral intermediate (tetralin hydroperoxide) is produced from an achiral substrate (tetralin) via an autocatalytic pathway where the feedback mechanism is expected to generate a state of broken chiral symmetry. In order to test the amplification capacity of this reaction a computer analysis of the kinetic scheme is performed. This simulation is derived from the known kinetic scheme of autoxidation and is validated by fitting the experimentally observed data of hydroperoxide evolution. Calculations show that this model allows powerful amplification of enantiomeric excess and a transient amplification of the optical rotation. It is also demonstrated that the model system exhibits pronounced sensitivity toward any loss of absolute configuration of the involved chiral species. Since an amplification effect results exclusively at a high degree of stereoselectivity, it is concluded that stereospecific autocatalysis is possible in systems which show template reactions, crystallization, or colloidal effects. © 1993 Wiley-Liss, Inc.     

Stereoselective binding and activity of oxotremorine analogs at muscarinic receptors in rat brain.

The activities of the enantiomers of BM-5 were examined to measure muscarinic cholinergic selectivity in the central nervous system. Autoradiographic studies assessed the ability of each enantiomer to inhibit the binding of [3H]-(R)-quinuclidinyl benzilate ([3H]-(R)-QNB) to muscarinic receptors in the rat brain. (+)-(R)-BM-5 inhibited [3H]-(R)-QNB binding to rat brain sections at concentrations below 1.0 microM, while 100-fold higher concentrations of (-)-(S)-BM-5 were required for comparable levels of inhibition. Analysis of the autoradiograms indicated that both stereoisomers had a similar distribution of high affinity binding sites. Each enantiomer displayed higher affinity for muscarinic receptors in the superior colliculi and lower affinity for receptors in the cerebral cortex and hippocampus. (+)-(R)-BM-5 and oxotremorine inhibited adenylyl cyclase activity in the cerebral cortex with efficacies comparable to that for acetylcholine. (+)-(R)-BM-5 was 26-fold more potent than (-)-(S)-BM-5 in inhibiting adenylyl cyclase. Oxotremorine-M and carbamylcholine stimulated phosphoinositide turnover in the cerebral cortex. Oxotremorine had lower activity and (+)-(R)-BM-5 was essentially inactive at comparable concentrations. The difference in activity of the two enantiomers indicates a remarkable stereochemical selectivity for muscarinic receptors. The stereoselectivity index is comparable for both the autoradiographic assays (48) and measures of adenylyl cyclase activity (26) in the cerebral cortex.     

Lipid-dependent sequential allosteric activation of heat-sensing TRPV1 channels by anchor-stereoselective “hot” vanilloid compounds and analogs

Both a silent resident phosphatidylinositol lipid and a “hot” vanilloid agonist capsaicin or resiniferatoxin have been shown to share the same inter-subunit binding pocket between a voltage sensor like domain and a pore domain in TRPV1. However, how the vanilloid competes off the resident lipid for allosteric TRPV1 activation is unknown. Here, the in silico research suggested that anchor-stereoselective sequential cooperativity between an initial recessive transient silent weak ligand binding site and a subsequent dominant steady-state strong ligand binding site in the vanilloid pocket may facilitate the lipid release for allosteric activation of TRPV1 by vanilloids or analogs upon non-covalent interactions. Thus, the resident lipid may play a critical role in allosteric activation of TRPV1 by vanilloid compounds and analogs.     

Stereoselective Accumulation of Propranolol Enantiomers in K562 and K562/ADR Cells

The stereoselective uptake of propranolol enantiomers was investigated by using the K562 and K562 adriamycin‐resistant cell line (K562/ADR) as a model. An enantioselective RP‐HPLC method was applied to determine the accumulation of propranolol (PPL) stereoisomers in K562 and K562/ADR cells. The concentration, time and temperature dependent studies showed that the accumulation of S‐(?)‐PPL was higher than R‐(+)‐PPL in K562 cells and uptake of R‐(+)‐PPL was significantly higher than that of S‐(?)‐PPL in K562/ADR cells. The results indicate the enantioselective accumulation of propranolol enantiomers in K562 and K562 / ADR cells. Chirality 25:361–364, 2013. © 2013 Wiley Periodicals, Inc.     

羊毛硫肽类化合物(Lanthipeptide)生物合成新进展

羊毛硫肽化合物(Lanthipeptides)是由核糖体合成并经过翻译后修饰得到的一大类肽类天然产物。这类化合物广泛的产生于不同种类的细菌,具有丰富的结构和生物活性多样性,为活性药物研究和开发提供重要的来源。本文综述了近几年来羊毛硫肽化合物生物合成进展,从其合成酶结构,进化机制,区域和立体选择性控制等方面进行了简要的讨论,展示了羊毛硫肽类化合物生物合成中特殊而迷人的酶学机制。