Synthesis of all four stereoisomers of 3-amino-2-hydroxybutanoic acids

All four stereoisomers of 3-amino-2-hydroxybutanoic acids were been obtained as single enantiomers via stereospecific reactions from D-gulonic acid gamma-lactone and D-glucono-delta-lactone.     

Synthesis of four stereoisomers of 1,4-thiazane-3-carboxylic acid 1-oxide via the asymmetric transformation (combined isomerization-preferential crystallization) of 1,4-thiazane-3-carboxylic acid

In order to synthesize four stereoisomers of 1,4-thiazane-3-carboxylic acid 1-oxide (TCA SO), (S)-1,4thiazane-3-carboxylic acid [(S)-TCA], which is one of the precursors, was prepared by the asymmetric transformation (combined isomerization-preferential crystallization) of (RS)-TCA. This asymmetric transformation was used (2R, 3R)-tartaric acid [(R)-TA] as a resolving agent and salicylaldehyde as the epimerization catalyst in propanoic acid at 110 degrees C to afford a salt of (S)-TCA with (R)-TA in 100% de with a yield of over 90%. Optically pure (S)-TCA was obtained by treating the salt with triethylamine in methanol in a yield of over 80%, based on (RS)-TCA as the starting material. In addition, asymmetric transformation of (R)-TCA gave (S)-TCA in a yield of 60-70%. (S)-TCA was oxidized by hydrogen peroxide in dilute hydrochloric acid to selectively crystallize (1S, 3S)-TCA.SO. (1R, 3S)-TCA SO of 70% de from the filtrate was allowed to form a salt with (R)-TA after a treatment with triethylamine to give (1R, 3S)-TCA SO as a single diastereoisomer. (1R, 3R)- and (1S, 3R)-TCA.SO were also prepared by starting from (R)-TCA that had been synthesized from L-cysteine.     

Synthesis of L-2-oxothiazolidine-4-carboxylic acid

An improved synthesis of L-2- oxothiazolidine -4-carboxylic acid is described. The new procedure, which leads to excellent yields of product, does not require the use of phosgene. The new method is thus less hazardous than the original one, and is readily adaptable to the preparation of 35S-labeled product.     

Inactivation of gamma-aminobutyric acid aminotransferase by (Z)-4-amino-2-fluorobut-2-enoic acid

(Z)-4-Amino-2-fluorobut-2-enoic acid (1) is shown to be a mechanism-based inactivator of pig brain gamma-aminobutyric acid aminotransferase. Approximately 750 inactivator molecules are consumed prior to complete enzyme inactivation. Concurrent with enzyme inactivation is the release of 708 +/- 79 fluoride ions; transamination occurs 737 +/- 15 times per inactivation event. Inactivation of [3H]pyridoxal 5'-phosphate ([3H]PLP) reconstituted GABA aminotransferase by 1 followed by denaturation releases [3H]PMP with no radioactivity remaining attached to the protein. A similar experiment carried out with 4-amino-5-fluoropent-2-enoic acid [Silverman, R. B., Invergo, B. J., & Mathew, J. (1986) J. Med. Chem. 29, 1840-1846] as the inactivator produces no [3H]PMP; rather, another radioactive species is released. These results support an inactivation mechanism for 1 that involves normal catalytic isomerization followed by active site nucleophilic attack on the activated Michael acceptor. A general hypothesis for predicting the inactivation mechanism (Michael addition vs enamine addition) of GABA aminotransferase inactivators is proposed.     

Synthesis and structure-activity relationship of 7-(substituted)-aminomethyl-4-quinolone-3-carboxylic acid derivatives

Gram-positive organisms have re-emerged as the major hospital pathogens, which make the unmet medical needs for antibacterial therapy even worse. In searching for potent agents against Gram-positive pathogens, novel 7-(substituted)-aminomethyl-quinolone-3-carboxylic acids were designed, synthesized, and evaluated for their antibacterial activities in vitro. Many 7-monoarylaminomethyl derivatives exhibited high potency against Gram-positive organisms compared to reference agents: vancomycin and pazufloxacin. Additionally, a few 7-monoalkylaminomethyl derivatives exhibited good activities against both Gram-positive and Gram-negative organisms.     

Biosynthesis of the 2-(aminomethyl)-4-(hydroxymethyl)furan subunit of methanofuran

2H- and 13C-labeled precursors were used to establish the pathway for the biosynthesis of the 2-(aminomethyl)-4-(hydroxymethyl)furan (F1) component of methanofuran in methanogenic archaebacteria. The extent and position of the label incorporated into F1 were measured from the mass spectrum of the diacetyl derivative of F1. [1,2-13C2]Acetate was found to be incorporated into two separate positions of the F1 molecule as a unit. The extent of incorporation of 13C2 into each of these positions was the same as that observed for the incorporation of acetate into the alanine and proline produced by the cells. From [2,2,2-2H3]acetate, deuterium was incorporated into two separate sites of the F1 molecule, one containing up to two deuteriums and the other only one. On the basis of the fragmentation pattern of the F1 diacetyl derivative, it was determined that two deuteriums were incorporated into the hydroxymethyl group at C-4 and one was incorporated at C-3 of the furan ring. The extent and distribution of the incorporated deuterium at the C-4 methylene were the same as that observed for C-6 of the glucose produced by the cells. On the basis of this and additional information presented in this paper, it is concluded that F1 is generated by the condensation of dihydroxyacetone phosphate with pyruvate. The resulting dihydroxy-substituted tetrahydrofuran after elimination of 2 mol of water would produce the phosphate ester of 2-carboxy-4-(hydroxymethyl)furan. Reduction of the carboxylic acid to an aldehyde and subsequent transamination would produce the phosphate ester of F1.     

Syntheses of (Z)-and (E)-4-amino-2-(trifluoromethyl)-2-butenoic acid and their inactivation of gamma-aminobutyric acid aminotransferase.

(Z)- and (E)-4-amino-2-(trifluoromethyl)-2-butenoic acid (4 and 5, respectively) were synthesized and investigated as potential mechanism-based inactivators of gamma-aminobutyric acid aminotransferase (GABA-AT) in a continuing effort to map the active site of this enzyme. The core alpha-trifluoromethyl-alpha,beta-unsaturated ester moiety was prepared via a Reformatsky/reductive elimination coupling of the key intermediates tert-butyl 2,2-dichloro-3,3,3-trifluoropropionate and N,N-bis(tert-butoxy-carbonyl)glycinal. Both 4 and 5 inhibited GABA-AT in a time-dependent manner, but displayed non-pseudo-first-order inactivation kinetics; initially, the inactivation rate increased with time. Further investigation demonstrated that the actual inactivator is generated enzymatically from 4 or 5. This inactivating species is released from the active site prior to inactivation, and as a result, 4 and 5 cannot be defined as mechanism-based inactivators. Furthermore, 4 and 5 are alternate substrates for GABA-AT, transaminated by the enzyme with Km values of 0.74 and 20.5 mM, respectively. Transamination occurs approximately 276 and 305 times per inactivation event for 4 and 5, respectively. The enzyme also catalyzes the elimination of the fluoride ion from 4 and 5. A mechanism to account for these observations is proposed.     

The effect of L-2-amino-4-methoxy-trans-3-butenoic acid on serine hydroxymethyl transferase

The tumour growth inhibitor L-2-amino-4-methoxy-trans-3-butenoic acid (Ro07-7957) inhibits serine hydroxymethyltransferase in cytosolic extracts of Walker carcinoma non-competitively with respect to L-serine with an apparent inhibition constant similar to the K_m-value for L-serine. The kinetics of inactivation suggest that it reacts as an irreversible substrate analogue. Incubation of Walker cells with Ro07-7957 causes an increase in serine hydroxymethyltransferase activity which is most pronounced at concentration ≤LD₅₀. This increase in enzyme activity does not occur in the presence of cycloheximide. These results suggest that inhibition of serine hydroxymethyltransferase in intact cells is accompanied by an increase in enzyme biosynthesis and that the growth inhibitory property of Ro07-7957 does not involve interference with the conversion of serine to glycine.     

Synthesis and biofilm inhibition studies of 2-(2-amino-6-arylpyrimidin-4-yl)quinazolin-4(3H)-ones

Synthesis of novel 4(3H)-quinazolinonyl aminopyrimidine derivatives has been achieved via quinazolinonyl enones which in turn were obtained from 2-acyl-4(3H)-quinazolinone. They have been assayed for biofilm inhibition against Gram-positive (methicillin-resistant Staphylococcus aureus (MRSA)) and Gram-negative bacteria (Acinetobacter baumannii). The analogues with 2,4,6-trimethoxy phenyl, 4-methylthio phenyl, and 3-bromo phenyl substituents (5h, 5j & 5k) have been shown to inhibit biofilm formation efficiently in MRSA with IC₅₀ values of 20.7–22.4 μM). The analogues 5h and 5j have demonstrated low toxicity in human cells in vitro and can be investigated further as leads.     

Enantioselective synthesis of (S)-2-amino-4-phenylbutanoic acid by the hydantoinase method

Biosynthesis of (S)-(+)-2-amino-4-phenylbutanoic acid (1) was performed by nonenantioselective hydantoinase and L-N-carbamoylase using racemic 5-[2-phenylethyl]-imidazolidine-2,4-dione (rac-2) as a substrate. The compounds involved in this biocatalysis process could be simultaneously resolved by high-performance liquid chromatography using Chirobiotic T column with a mobile phase of EtOH/H(2)O = 10/90 at pH 4.2-4.5. To our knowledge, this is the first report of the successful production of 1 by the combination of recombinant hydantoinase and L-N-carbamoylase.     

Stereoselective intramolecular cyclization to 4-(hydroxymethyl)-3-(1H-indolyl)oxazolidin-2-ones

A simple high-yield three-steps route to optically active 4-hydroxymethyl-3-(1H-indolyl)oxazolidin-2-ones from (S)-glycidol is described. The key intermediates (R)-oxiran-2-ylmethyl 1H-indol-4/-5-ylcarbamates are obtained in high yields from (S)-glycidol. These are readily transformed to oxazolidin-2-ones, very interesting building blocks in drug synthesis.     

Occurrence of stereoisomers of 1-(2'-pyrrolidinethione-3'-yl)- 1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid in fermented radish roots and their different mutagenic properties

Stereoisomers of the tetrahydro-beta-carboline derivative, 1-(2-pyrrolidinethione)-3-yl)-1,2,3,4-tetrahydro-beta-carboline- 3-carboxylic acid (PTCC), were formed from L-tryptophan with 4-methylthio-3-butenyl isothiocyanate, and their mutagenic properties and contents in different types of the radish products were studied. The isomers were identified as (1S*, 3S*, 3R*)- and (1R*, 3S*, 3R*)-PTCCs; the former was found as the major compound but had no mutagenic activity, while the latter was mutagenic toward Salmonella typhimurium TA 98 in the presence of a rat microsomal fraction. Both (1S*, 3S*, 3R*)- and (1R*, 3S*, 3R*)-PTCC were detected in a ratio of about 4:1 in a product fermented for 8 months, but only a trace was apparent in products manufactured within a few weeks.     

Synthesis and evaluation of [(1R)-1-amino-2-(2,5-difluorophenyl)ethyl]cyclohexanes and 4-[(1R)-1-amino-2-(2,5-difluorophenyl)ethyl]piperidines as DPP-4 inhibitors

A series of 4-amino cyclohexanes and 4-substituted piperidines were prepared and evaluated for inhibition of DPP-4. Analog 20q displayed both good DPP-4 potency and selectivity against other proteases, while derivative 20k displayed long half life and modest oral bioavailability in rat. The most potent analog, 3-(5-aminocarbonylpyridyl piperidine 53j, displayed excellent DPP-4 activity with good selectivity versus other proline enzymes.