Lactones 29. Enzymatic resolution of racemic γ-lactones

Studies on the application of commercially available enzymes to resolution of the racemic unsaturated γ-lactones: 5-(3-methylbutylidene)-4-methyl-tetrahydrofuran-2-one (1a) and 5-(3,3-dimethylbutylidene)-4-methyl-tetrahydrofuran-2-one (2a) are presented. Lipase PS, Rhizopus niveus lipase, Rhizopus arrhizus lipase, porcine pancreas lipase and hog liver esterase transformed substrates into their respective γ-keto acids with good efficiency (50–75%). Three of them hydrolysed the studied lactones with moderate enantioselectivity. Enantiomeric excesses determined by GC for the unreacted lactones were in the range of 20–60%. Lipase PS preferentially hydrolysed the (+) enantiomers of lactones 1a and 2a whereas R. niveus lipase hydrolysed the (?) enantiomers, respectively.     

Regioselective acylation of several polyhydroxylated natural compounds by Candida antarctica lipase B

Regioselective acylation of four polyhydroxylated natural compounds, deacetyl asperulosidic acid (1), asperulosidic acid (2), puerarin (3) and resveratrol (4) by Candida antarctica Lipase B in the presence of various acyl donors (vinyl acetate, vinyl decanoate or vinyl cinnamoate) was studied. Compounds 1, 2 and 4 were regioselectively acetylated with vinyl acetate to afford products, 3'-O-acetyl-10-O-deacetylasperulosidic acid (1a), 3',6'-O-diacetyl-10-O-deacetylasperulosidic acid (1b), 3'-O-acetylasperulosidic acid (2a), 3',6'-O-diacetylasperulosidic acid (2b), 4'-O-acetylresveratrol (4a), respectively, with yields of 22 to 50%, while reactions with vinyl decanoate and vinyl cinnamoate were slow with lower yields. Compound 3 was readily acylated with all three acyl donors and quantitatively converted to products 6'-O-acetylpuerarin (3a), 6'-O-decanoylpuerarin (3b), 6'-O-cinnamoylpuerarin (3c), respectively. The structures of these acylated products were determined by spectroscopic methods (MS and NMR).     

Temperature-dependence of enantioselectivity and desymmetrization in the acetylation of 2-mono- and 2,2-di-substituted 1,3-propanediols by a novel lipase isolated from the yeast Cryptococcus spp. S-2

The effect of temperature on enantioselectivity and desymmetrization in the acetylation of 2-phenyl-1,3-propanediol (1a), 2-benzyl-1,3-propanediol (1b), 2-methyl-2-phenyl-1,3-propanediol (1c) and 2-benzyl-2-methyl-1,3-propanediol (1d) by a novel lipase (CSL) isolated from the yeast Cryptococcus spp. S-2 was studied. Desymmetrization of 1a, 1c and 1d by CSL-catalyzed acetylation was observed in the temperature range of -20°C to 40°C, while diacetylation of 1b occurred considerably even at 0°C. The kinetic parameters of the selectivity indicated that the acetylation of 1a is an entropy controlled process whereas the reaction of 1c and 1d is mainly controlled by the enthalpy term. In the monoacetylation of the diol 1d, the preferred configuration in the enantiomeric induction by CSL was opposite to that of immobilized porcine pancreatic lipase (PPL).     

Stereochemistry of the Enzymatic Reduction of 2-(4-Methoxybenzyl)-1-Cyclohexanone by Solanum Aviculare Cells in vitro

During the investigation of chemical properties of the dicyclic system of insect juvenile hormone analogues, biotransformation of 2-(4-methoxybenzyl)-1-cyclohexanone (1) by plant cell cultures was studied. Among other components, the cis-(1S, 2S)- and cis-(1R, 2R)-2-(4-methoxybenzyl)-1-cyclohexanol enantiomers 2a and 2b were found in the reaction mixture. Higher stereoselectivity of the biotransformation was observed for trans-(1S, 2R)-enantiomer 3a formation, which occurred in at least 60% of calculated enantiomeric excess.     

Biotransformation of podophyllotoxin by Hordeum vulgare cell suspension cultures

Hordeum vulgare cell suspension cultures were used to modify podophyllotoxin (1) One major product (1a) and one minor product (1b) were detected in both the culture medium and cells. To optimize the yield of compound 1a, we showed that: (1) the optimal concentration of added podophyllotoxin (1) was 33 mg L^-1; higher concentrations caused cell toxicity; (2) the stage of the cell cycle (lag/log/stationary) at which podophyllotoxin was added only marginally affected the yield of compound 1a; the optimal addition time was after lag phase, in which the yield of compound 1a reached ca. 76%, and (3) biotransformation of podophyllotoxin (1) was relatively slow; podophyllotoxin fed at 4 days after subculture resulted in yields of compound 1a of ca. 56, 64 and 76% after an additional 3, 6 and 10 days of incubation, respectively. Product 1a was purified and identified as isopicropodophyllone (1a) based on MS and NMR data.     

An Efficient Chemoenzymatic Synthesis of S-(-)-Fenpropimorph

First enantioselective synthesis of S-(-)-1-[3-(4-tert-butylphenyl)-2-methyl]propyl-cis-3,5-dimethylmorpholine (6), biologically active enantiomer of the systematic fungicide fenpropimorph, is reported. It comprises reacting 4-tert-butylbenzylbromide with methyldiethylmalonate, decarbethoxylation of 2 into racemic 3-(4-tert-butylphenyl)-2-methylpropionic acid ethylester (3) in DMSO in the presence of alkali, then Pseudomonas sp. lipase catalyzed kinetic resolution of racemic 3 into S-(+)-acid (4), base-catalyzed racemization and recycling of the R-(-)-ester 3, acylation of cis-3,5-dimethylmorpholine, and final reduction of the intermediary amide 5 to provide enantiomerically pure S-(-)-6.     

Biotransformation of γ-terpinene using Stemphylium botryosum (Wallroth) yields p-mentha-1,4-dien-9-ol, a novel odorous monoterpenol

A wild strain of Stemphylium botryosum, when grown submerged in the presence of γ-terpinene (1), yielded the novel highly odour active terpene alcohol (2), whose structure was established by spectroscopic means as p-mentha-1,4-dien-9-ol. During cultivation (2) was further oxidized, predominantly to the corresponding aromatic alcohol p-cymene-9-ol (5). The enantioselective enzymatic introduction of the hydroxyl group at the non-activated C9 of (1) resulted in an enantiomeric excess (ee) of 74% for (2) and 70% for (5), respectively.     

Sol–gels and cross-linked aggregates of lipase PS from Burkholderia cepacia and their application in dry organic solvents

Lipase PS from Burkholderia cepacia (formerly Pseudomonas cepacia) was successfully immobilized in sol–gels under low methanol conditions using lyophilization in order to dry the gel. The enzyme was also cross-linked with glutaraldehyde to CLEAs without any additives. These immobilized enzyme preparations were employed for the highly enantioselective acylations of 1-phenylethanol (1), 1-(2-furyl)ethanol (2) and N-acylated 1-amino-2-phenylethanol (3) with vinyl acetate in organic solvents. Enzymatic hydrolysis of the obtained ester product was observed as a side reaction of the acylation of 3 in the presence of lipase PS powder. Hydrolysis was suppressed when the immobilized preparations of lipase PS were used.     

R( − )pantoyllactone-β-d-glucopyranoside: Characterization of a metabolite from rice seedlings

A new derivative of pantoic acid, R( - ) pantoyllactone-β-d glucopyranoside, has been isolated from rice seedlings and its structure determined. β-Glucosidase hydrolysed it to -glucose and R( - )pantoyllactone. Alkaline hydrolysis converted it to the salt of 2-R( - )pantoic acid β- -glucopyranoside. It accumulated in rice shoots but not in roots.     

Lipase-catalyzed kinetic resolutions of racemic β- and γ-thiolactones

Several racemic β- and γ-thiolactones were synthesized and kinetic resolutions of them were executed using lipases. While a lipase from Pseudomonas cepacia (PCL) showed the highest enantioselectivity for (S)-form (>99% ee_S at 53% conversion, E > 100) in the kinetic resolution of racemic -methyl-β-propiothiolactone (rac-MPTL), it showed no hydrolysis activity in the kinetic resolution of -benzyl--methyl-β-propiothiolactone (rac-BMPTL), suggesting that the changes in the size of alkyl group from rac-MPTL to rac-BMPTL leads to lower hydrolysis activity and enantioselectivity. In contrast, racemic γ-butyrothiolactones were hydrolyzed by several lipases with low enantioselectivity, whereas a lipase from Candida antarctica (CAL) showed moderate enantioselectivity for (S)-form (>99% ee_S at 76% conversion, E = 11) in the kinetic resolution of racemic -methyl-γ-butyrothiolactone (rac-MBTL). Computer-aided molecular modeling was also performed to investigate the enantioselectivites and activities of PCL toward β-propiothiolactones. The computer modeling results suggest that the alkyl side chains of β-propiothiolactones and γ-butyrothiolactones interact with amino acid residues around hydrophobic crevice, which affects the activity of PCL.     

2-substituted-3H-indol-3-one-1-oxides: preparation and radical trapping properties

A series of 2-alkyl and 2-aryl substituted-3H-indol-3-one-1-oxides was prepared and evaluated for its radical trapping properties. Spin trapping and electron paramagnetic resonance experiments demonstrate the ability of these indolone-1-oxides to trap hetero- and carbon-centered radicals. The most stable spin adducts (lifetime of several hours) are obtained with 2-alkyl substituted nitrones, the 2-ethyl-5,6-dioxolo-3H-indolone-1-oxide, 5e and the 2-secbutyl-3H-indolone-1-oxide, 5f. These two nitrones are also sensitive to redox reactions in solution. Therefore this indolone-1-oxide series lacking a β-hydrogen atom gives rise to highly stable adducts with free radicals.     

The biotransformation of methyl ent-15-oxokaur-16-en-19-oate by Rhizopus stolonifer and Mucor plumbeus

Incubation of methyl ent-15-oxokaur-16-en-19-oate with Rhizopus stolonifer and Mucor plumbeus gave methyl ent-7β,11-dihydroxy-15-oxokauran-19-oate and methyl ent-7β,16β-dihydroxy-15-oxokauran-19-oate, respectively.     

Book Review

LEGGETT, W. F. Ancient and Medieval Dyes. 5 × 8 in. 96 pp. Cloth. Chemical Publishing Co., Brooklyn, N. Y. 1944. $2.25.

Microtechnic In General. McCARTNEY, J. E. A new immersion oil “polyric”. J. Path. & Bact., 56, 265-6. 1944.

NICKERSON, MARK. A dry ice freezing unit for rotary microtomes. Science, 100, 177-8. 1944.

Dyes And Thedx Biological Uses. BERGEIM, FRANK H., and BRAKER, WILLIAM. Homosulfanilamides. J. Amer. Chem. Soc., 66, 1459. 1944.

CALDWELL, W. T., TYSON, F. T., and LAUER, LOTHAR. Substituted 2-sulfonamido-5-aminopyridines. II. J. Amer. Chem. Soc., 66, 1479. 1944.

JOHNS, C. K. Dye concentration in resazurin tablets. Amer. J. Pub. Health, 34, 955-8. 1944.

SMITH, WINSLOW WHITNEY. Relative sensitivity of different phases of growth curve of Bact. salmonicida to alkaline acriflavine. Proc. Soc. Exp. Biol. & Med., 56, 240-2. 1844.

VAN ARENDONK, A. M., and SHOULE, H. A. Dialkylaminoalkyl derivatives of substituted quinolines and quinaldines. J. Amer. Chem. Soc., 66, 1284. 1944.

WHEELER, KEITH, and DEGERING, E. F. Preparation and properties of certain derivatives of sulfamide. J. Amer. Chem. Soc., 66, 1242. 1944.

Animal Microtechnic. BOARDMAN, EDWARD T. Methods for collecting ticks for study and delineation. J. Parasitology, 30, 57-9. 1944.

DICKIE, MARGARET M. A new differential stain for mouse pituitary. Science, 100, 297-8. 1944.

GOVAN, A. D. TELFORD. Fat staining by Sudan dyes suspended in watery media. J. Path. & Bact., 56, 262-4. 1944.

LILLIE, R. D., and ASHBURN, L. L. Supersaturated solutions of fat stains in dilute isopropanol for demonstration of acute fatty degenerations not shown by Herxheimer technic. Arch. Path., 36, 432. 1943.

MULLEN, J. P. A convenient and rapid method for staining glycogen in paraffin sections with Best's carmine stain. Amer. J. Clin. Path., Tech. Sect., 8, 9-10. 1944.

NYKA, W. A method for staining the rickettsiae of typhns in histological sections. J. Path. & Bact., 56, 264. 1944.

POPPER, HANS, GYORGY, PAUL, and GOLDBLATT, H. Fluorescent material (ceroid) in experimental nutritional cirrhosis. Arch. Path., 37, 161. 1944.

SMALL, C. S., and SCHULTZ, M. A. Sustaining faded tissue sections. Amer. J. Clin. Path., Tech. Sect., 7, 66-7. 1943.

YOFFEY, J. M., and PARNELL, J. The lymphocyte content of rabbit bone marrow. J. Anat., 78, 109-12. 1944.

ZIEGLER, E. E. Hematoxylin-eosin tissue stain. An improved, rapid, and uniform technic. Arch. Path., 37, 68. 1044.

Plant Microtechnic. HAASIS, FERDINAND W. Staining rubber in ground or milled plant tissues. Ind. and Eng. Chem., Anal. Ed., 16, 480. 1944.

PARRIS, G. K. A simple nuclear stain and staining technique for Helminthosporia. Phytopathology, 34, 700. 1944.

Microorganisms. DARZINS, E. Rickettsienstudien. Zentbl. Bakl., Abt. I, Orig., 151, 18-20. 1943.

GOHAR, M. A. A staining method for Corynebacterium diphtheriae. J. Bact., 47, 575. 1944.

GRAY, P. H. H. Two-stain method for direct bacteria count. J. Milk Techn., 6, 76. 1943.     

Preparation of both enantiomers of methyl 3-benzoyloxypentanoate by enzyme-catalysed hydrolysis of corresponding racemic nitrile and amide

Rhodococcus rhodochrous IFO 15564 enantioselectively hydrolysed racemic 3-benzoyloxypentanenitrile and 3-benzoyloxypentanamide to afford (R)-amide and (S)-car☐ylic acid with high enantiomeric excess (> 90%). In this reaction, both enantiomers of the starting nitrile were converted to the amide by nitrile hydratase, and amidase-catalysed enantioselective hydrolysis of the amide was responsible for the kinetic resolution. The lack of enantioselectivity of the nittile hydratase toward the racemic nitrile forms a marked contrast to the case of previously reported highly enantioselective conversion of prochiral 3-benzoyloxypentanedinitrile by this enzyme. since (R)-amide could be hydrolysed chemically to (R)-car☐ylic acid without any loss of its ee, the present microbial kinetic resolution serves as an effective method for preparing both enantiomers of synthetically useful 3-hydroxypentanoic acid derivatives.     

Influence of precursors and additives on microbial lipases stabilized by sol-gel entrapment

Sol-gel entrapment of microbial lipases from Candida cylindracea (Cc lipase), Pseudomonas fluorescens (Lipase AK), and Pseudomonas cepacia (Lipase PS), using as precursors tetraethoxysilane (TEOS) and silanes of type R-Si(OEt)₃ with alkyl or aryl R groups, has been investigated. Three different methods using these precursors were tried exhibiting protein immobilization yields in the range of 20-50%. Hydrolysis of emulsified olive oil, esterification of lauric acid with 1-octanol and enantioselective acylation of 2-pentanol have been used as model reactions for testing the properties of the encapsulated lipases. The recovery yields of the enzyme activity in the esterification reaction were between 20-68%, the best performance being achieved with phenyltriethoxysilane and tetraethoxysilane precursors at 3:1 molar ratio. When testing the entrapped Lipase AK in the enantioselective acylation reaction of 2-pentanol, activity recovery yields up to 32% related to the free enzyme were obtained and the immobilization increased the enantioselectivity of the enzyme.     

Alkaloids from Heimia salicifolia

Two alkaloids, 9β,2′-dihydroxy-4′′,5′′-dimethoxy-lythran-12-one or 9β-hydroxyvertine (1) and (2S,4S,10R)-4-(3-hydroxy-4-methoxyphenyl)-quinolizidin-2-acetate (2), as well as seven known alkaloids, lythrine (3), dehydrodecodine (4), lythridine (5), vertine (6), heimidine (7), lyfoline (8) and epi-lyfoline (9), were isolated from Heimia salicifolia. The structures of these compounds were elucidated by extensive spectroscopic techniques. Furthermore, the structures of 2, 3, and 6 were confirmed by X-ray crystallography, including absolute configuration determination of 2 and 6. Compounds 6 and 9 showed moderate antimalarial activity.     

紫陀螺菌化学成分及其抑制肿瘤细胞增殖活性的研究

以紫陀螺菌为对象,研究其子实体的化学成分及其抑制肿瘤细胞增殖活性。采用溶剂提取、柱层析和高效液相色谱等方法分离纯化化学成分,通过核磁共振和质谱技术鉴定单体化合物结构,运用结晶紫法评价单体化合物抑制肿瘤细胞增殖活性。从乙酸乙酯提取物中共分离鉴定6个单体化合物,分别为(22E,24R)-麦角甾-5,7,22-三烯-3β-醇(1)、3β,5α-二羟基-(22E,24R)-麦角甾-7,22-二烯-6-酮(2)、(22E,24R)-麦角甾-7,22-二烯-3β,5α,6β-三醇(3)、吲哚-3-甲酸甲酯(4)、4,4-二甲基-1,7-庚二酸(5)和(8E,10E)-12羰基十八碳-8,10-二烯酸(6),其中化合物1为主要成分,相对含量为23.8%。活性测试结果表明3对人乳腺癌细胞株MCF-7 细胞、人胰腺癌细胞株PANC-1细胞和人乳腺癌细胞株MDA-MB-231细胞具有微弱的细胞增殖抑制活性。本研究首次报道了紫陀螺菌化学成分,对深入挖掘其在健康领域中的开发价值具有重要意义。     

Production of malodorous steroids from androsta-5,16-dienes and androsta-4,16-dienes by Corynebacteria and other human axillary bacteria

The biotransformations of a number of steroids, chiefly 5,6,16,17-tetradehydro-androstanes, are reported. The strains investigated were Corynebacteria sp. G38, G40, G41, B, Brevis sp. CW5 and Micrococcus sp. M-DH2. Corynebacterium sp. G41 proved remarkably efficient in effecting oxidative isomerisation of 5-ene-3-sterols into the corresponding 4-en-3-ones. The main biochemical reactions involved were oxidation at C-3; no reduction processes were observed. Conversions of 3β-sterols into the C-3 oxo-steroids were high, but were correspondingly low for the 3-sterol epimers. Androsta-4,16-dien-3-one and 5β-androsta-16-en-3-one are crucial to the formation of malodour. The rate of formation of these compounds was measured over 72 h incubation periods using three substrates: androsta-5,16-dien-3β-ol, androsta-4,16-dien-3β-ol and androsta-5,16-dien-3-one. Induction studies of the transformation of the androsta-5,16-dien-3β-ol into the very odorous compound androsta-4,16-dien-3-one showed that cells incubated with a mixture of antibiotics displayed the same extent of biotransformation as normal cells if the concentration of antibiotic was low (1, 3, 5 and 7 μg/ml), although at concentrations higher than 10 μg/ml, biotransformation yields were reduced. Pre-incubation with a 3β-fluoro-steroid inhibited the formation of the odorous androsta-4,16-dien-3-one.     

Δ-β-Sitosten-3-one from β-sitosterol by leaf homogenates

β-Sitosterol-4-¹⁴C is metabolized to Δ⁴-β-sitosten-3-one by Cheiranthus cheiri leaf homogenates. Greater than 60% conversion occurs within 2 hr. Under identical conditions, leaf homogenates of Strophanthus kombé fail to metabolize β-sitosterol, while Digitalis purpurea leaf homogenates yield only very small amounts of the metabolite.     

Application of lipase-catalyzed transformations for the synthesis of insect pheromones and related compounds

This review describes efficient means of preparing optically pure insect pheromones and related compounds via lipase-catalyzed enantioselective reaction on a large scale. (1) A new synthesis of the Japanese beetle pheromone, (R,Z)-(−)-5-(1-decenyl)oxacyclopentan-2-one, established by a combination of two lipase-catalyzed transformation was demonstrated. (2) A chemico-enzymatic procedure for the syntheses of both enantiomers of cupreous chafer beetle pheromone, (R,Z)- and (S,Z)-5-(1-octenyl)oxacyclopentan-2-one, was described. (3) An optical resolution of (±)-2,3-epoxy-8-methyl-1-nonanol, the key intermediate of the synthesis of gypsy moth pheromone, was demonstrated. (4) A practical chemico-enzymatic synthesis of (+)-disparlure in large scale was demonstrated. (5) A facile synthesis of carboxyalkyl acrylate, which is special monomers in the synthesis of the new polymers, by two lipase-catalyzed regioselective reactions was described.