环氧丙醇酯立体专-性水解酶产生菌的筛选*

从土壤中筛选出一株能拆分（Ｒ,Ｓ）－环氧丙醇丁酸酯的根霉（Ｒｈｉｚｏｐｕｓｓｐ．Ｂｃ０－０９）,该菌株所产胞外脂肪酶在水解环氧丙醇丁酸酯的反应中具有良好的立体专一性。在ｐＨ恒定７．０的条件下,以其发酵液水解底物,当转化率为５８％时,残留的（Ｒ）－环氧丙醇丁酸酯的光学纯度（对映体过量值）达９６．１％。     

环氧丙醇酯立体专一性水解酶产生菌的筛选

从土壤中筛选出一株能拆分－环氧丙醇丁酸的根,该菌株所产胞外脂酶在水解环氧丙醇丁酯的反应中具有良好的立体专一性。     

米根霉发酵生产L-乳酸

报道了Ｌ－乳酸菌株的分离与筛选,探讨了不同碳源、氮源、通气量、温度等发酵条件对产Ｌ－乳酸的影响,从７８株米根霉中筛选出１３株产Ｌ－乳酸较高的菌株,其中米根霉（Ｒｈｉｚｏｐｕｓ ｏｒｙｚａｅ）Ｒｓ９２８产Ｌ－乳酸最高,产酸最稳定。试验结果表明,该菌株最适发酵培养组成（％）：淀粉水解糖１６,ＭｇＳＯ４ ０．０８,ＫＨ２ＰＯ４ ０．０５,ＺｎＳＯ４ ０．０１,ＣａＣＯ３ ７,ｐＨ自然。在６０ｔ发酵罐中,     

冬凌草中一新的二萜成分--冬凌草辛素

从河南省鹤壁地区产冬凌〖Ｉｓｏｄｏｎ ｒｕｂｅｓｃｅｎｓ（Ｈｅｍｓｌ．）Ｈａｒａ〗叶中分得一个新地二萜化合物,命名为冬凌草辛素（Ｒｕｂｅｓｃｅｎｓｉｎ Ｈ）〈经ＮＭＲ谱等确定其结构为６β,７β,１４β,１５Ｒ－四羟基－１１β－甲酰氧基－７α,２０－环氧－对映－贝壳杉－１６－烯。     

微生物酶不对称水解合成S（＋）—布洛芬的研究：Ⅱ.反应条件和产物提取

皮状丝孢酵母具有较强不对称水解底物专一性。在试验的五种布洛芬消旋酯中,水解甲酯和异丙酯生成Ｓ（＋）－布洛芬ｅｅ可达９７％,乙酯为９３％以上;而水解活性以乙酯最强。转化率高于３０５。不对称水解最适ｐＨ６．５－７．０;温度在２８－３７℃范围内拆分能力无明显差别。该酵母的水解酶为胞内酶,将酵母细胞制成丙酮干粉进行水解可提高立体专一性。产物Ｓ（＋）－布洛芬可借助于酸碱反应和有机溶剂提取得到,同时回收未水解     

青年男性高原移居者最大摄氧量计算公式的推导

最大摄氧量（Ｖｏ２ｍａｘ）是评价人体体力的重要指标,其测定方法分直接法和间接法两种。目前所推导的间接计算公式都是在平原、或是在进入高原初期推导的,不适用于高原习服人群。本研究采用逐步回归的方法,推导出移居高原７－２７个月、不同高度的青年男性Ｖｏ２ｍａｘ间接计算公式。在海拔３６８０ｍ地区,Ｖｏ２ｍａｘ（Ｌ／ｍｉｎ）＝１．１５３１＋０．００７３２７身高（ｃｍ）＋０．０１６１３体重（ｋｇ）－０．００５８８３晨脉（ｂ／ｍｉｎ）－０．００４５３４运动心率（６０Ｗ,６／ｍｉｎ）,Ｒ＝０．７４５,Ｐ＜０．０１,ＳＳ＝３．７７９９;或Ｖｏ２ｍａｘ（Ｌ／ｍｉｎ）＝１．２１８６＋０．０１９８４体重（ｋｇ）＋０．０７２５９肺活量（Ｌ）－０．００６６５９晨脉（ｂ／ｍｉｎ）,Ｒ＝０．７１３,ｐ＜０．０１,ｓｓ＝３．９６３６。在４３５０ｍ地区,Ｖｏ２．ｍａｘ（Ｌ／ｍｉｎ）＝０．４９１７＋０．０１６８７体重（ｋｇ）＋０．１１０９肺活量（Ｌ）＋０．００１９８３屏气时间（Ｓ）,Ｒ＝０．７８１,Ｐ＜０．０１,ＳＳ＝２．１３５６。计算值与实测值比较,变异系数在１３％以内,结果准确可靠,适用于青年男性高原习服移居者。     

幼狮心电图的测定和分析

本研究采用标准双极肢导联（Ｉ,Ⅱ,Ⅲ）和加压单极肢导联（ａＶＲ,ａＶＬ,ａＶＦ）,对未经麻醉并处于清醒安静状态下的４只全同胞幼狮进行了心电图测定。结果表明,幼狮的平均心率为１５９±１次／ｍｉｎ,均为窦性心律,ＱＲＳ波平均心电轴为６８±１８°。Ｐ波,ＱＲＳ综合波及Ｔ波持续时间分别为０．０５５±０．００４ｓ,０．０６７±０．００６ｓ和０．０６９±０．０１２ｓ。Ｒ－Ｒ,Ｐ－Ｒ,Ｓ－Ｔ及Ｑ－Ｔ间期分别为０．３７８±０．００２ｓ,０．０８２±０．００６ｓ,０．０７８±０．００５８和０．２２５±０．０１７ｓ。Ｐ波的形态在Ｉ,Ⅱ,Ⅲ和ａＶＦ导联中呈正向,而在ａＶＲ及ａＶＬ导联中呈负向;ＱＲＳ综合波的波形呈现一定的规律性,在Ｉ,Ⅱ,Ⅲ及ａＶＦ导联中主要表现为Ｒ型正向波,而在ａＶＲ,ａＶＬ导联中则表现为Ｑ或Ｓ型负向波;Ｓ－Ｔ段无出现移位现象;Ｔ波在ａＶＬ导联中,没有表现出明显的规律性,而在Ｉ,Ⅱ,Ⅲ和ａＶＦ导联中,主要呈正向,在ａＶＲ和ａＶＬ导联中则往往呈负向．     

ras基因和表皮生长因子受体在胃癌中的表达及其病理临床意义

应用免疫组化方法检测ｒａｓ基因蛋白和表皮生长因子受体（ＥＧＦＲ）在７５例胃癌组织中的表达,研究它们与胃癌病理特征及预后关系。７５例胃癌ｒａｓ基因表达阳性率为４６．７％,与胃癌的分化程度,生长方式,浸润深度和淋巴转移呈明显正相关（Ｐ＜０．０５）;ＥＧＦＲ表达阳性率６１．３％,癌旁组织及新生血管有阳性表达;ＥＧＦＲ表达与胃癌大体类型,分化程度,生长方式和淋巴结转移呈正相关（Ｐ＜０．０５）;ｒａｓ蛋白表达与ＥＧＦＲ表达有明显关系（Ｐ＜０．０５）;Ｋａｐｌａｎ－Ｍｅｉｅｒ生存分析ｒａｓ蛋白和ＥＧＦＲ表达与胃癌的预后有明显的关系（Ｐ＜０．０１）。两者表达的检测有助于判断胃癌的恶性程度和预后估计。     

急性低氧诱导大鼠皮层和海马区自由基增殖

实验模型采用麻醉大鼠人工吸入１０．５％－１１．０％氮氧混合气体（相当于５０００ｍ海拔高度），于低氧前５ｍｉｎ由股静脉注入α－苯基－Ｎ－叔丁基甲亚胺－Ｎ－氧化物（ＰＢＮ，α－ｐｈｅｎｙｌ－ｔｅｒ－ｂｕｔｙｌｎｉ－ｔｒｏｎｅ）。结果发现低氧１５ｍｉｎ后，皮层和海马ＰＢＮ－自旋加合物电子自旋共振（ＥＳＲ，ｅｌｅｃｔｒｏｎ ｓｐｉｎ ｒｅｓｏｎａｎｃｅ）信号强度显著大于常氧对照组（ｎ＝５，Ｐ〈０．０１）。     

8%—9%和12%—13%低氧气体诱导大鼠左右心室肌自由基增殖

α－苯基－Ｎ－叔丁基甲亚胺－Ｎ－氧化物（ＰＢＮ,α－ｐｈｅｎｙｌ－ｔｅｒｔ－ｂｕｔｙｌｎｉｔｒｏｎｅ）由股静脉注入ＳＤ大鼠体内,鼠暴露于８％－９％Ｏ２（ｉｎＮ２）１５ｍｉｎ,左心肌ＰＢＮ－自旋加合物电子自旋共振（ＥＳＲ,ｅｌｅｃｔｒｏｎｓｐｉｎ ｒｅｓｏｎａｎｃｅ）信号明显增强（与常氧组比,ｎ＝５,Ｐ〈０．０５）。,而右心肌变化不明显（ｎ＝５,Ｐ〉０．０５）。暴露于１２％－１３％Ｏ２（ｉｎＮ２）的     

Efficient and selective microbial esterification with dry mycelium of Rhizopus oryzae

The use of dry mycelium of Rhizopus oryzae as biocatalyst for ester production in organic solvent has been studied. Mycelia with notable carboxylesterase activity were produced when different Tweens (20, 40, 60 and 80) were employed as main carbon source for the growth. Dry mycelium of four strains of Rhizopus oryzae proved effective for efficiently catalysing the synthesis of different flavour esters (hexylacetate and butyrate, geranylacetate and butyrate) starting from the corresponding alcohol and free acid, including acetic acid. The esterification of the racemic mixture of 2-octanol and butyric acid proceeded with high enantioselectivity (R-ester produced with enantiomeric excess > or =97%) when Rhizopus oryzae CBS 112.07 and Rhizopus oryzae CBS 260.28 were employed.     

Chiral synthesis of alpha-phenylpyridylmethanols with Rhizopus arrhizus

The bioreduction of 2-benzoylpyridine (1a) with Rhizopus arrhizus afforded (S)-(+)-alpha-phenyl-2-pyridylmethanol (2a) in 82% enantiomeric excess (e.e.) while the asymmetric hydrolysis of its racemic acetate resulted in the antipode (R)-(-)-2a with 24% optical purity.     

Newly isolated Streptomyces spp. as enantioselective biocatalysts: hydrolysis of 1,2-O-isopropylidene glycerol racemic esters

AIMS: To identify microbial strains with esterase activity able to enantioselectively hydrolyse esters of (R,S)-1,2-O-isopropylidene glycerol. METHODS AND RESULTS: The microbial hydrolysis of various racemic esters of 1,2-O-isopropylidene glycerol (IPG) was attempted by screening among Streptomyces spp. previously selected on the basis of their carboxylesterase activity. The best results were observed in the hydrolysis of butyrate ester and two strains appeared promising as they showed opposite enantioselectivity: Streptomyces sp. 90852 gave predominantly (S)-IPG, while strain 90930 mostly gave the R-alcohol. Streptomyces sp. 90930 was identified as Streptomyces violaceusniger, whereas Streptomyces sp. 90852 is a new species belonging to the Streptomyces violaceus taxon. The carboxylesterase belonging to strain 90852 gave a maximum value of enantiomeric ratio (E) of 14-16. This strain was lyophilized and used as dry mycelium for catalysing the synthesis of isopropylidene glycerol butyrate in heptane showing reaction rate and enantioselectivity (E = 6.6) lower than what observed for the hydrolysis. CONCLUSIONS: A new esterase with enantioselective activity towards (R,S)-IPG butyrate has been selected. The best enantioselectivity is similar or even better than the highest reported value in the literature with commercial enzymes. The enzyme is produced by a new species belonging to the S. violaceus taxon. SIGNIFICANCE AND IMPACT OF THE STUDY: New esterases from streptomycetes can be employed for the enantioselective hydrolysis of chiral esters derived from primary alcohols, not efficiently resolved with commercial enzymes.     

Microbial production of optically active beta-phenylalanine through stereoselective degradation of racemic beta-phenylalanine

The ability to produce (R)- or (S)-beta-phenylalanine from racemic beta-phenylalanine through stereoselective degradation was screened for. Variovorax sp. JH2 and Arthrobacter sp. the faculty of Agriculture, Kyoto University (AKU) 638 were found to be potential catalysts for (R)- and (S)-beta-phenylalanine production respectively. On 192 h cultivation of Variovorax sp. in medium containing 1.0% (w/v) racemic beta-phenylalanine, 0.46% (w/v) (R)-beta-phenylalanine with an enantiomeric purity of 99% e.e. was obtained. The initial step of the (S)-isomer degradation was stereoselective transamination. On 312 h cultivation of Arthrobacter sp. in medium containing 1.0% (w/v) racemic beta-phenylalanine, 0.51% (w/v) (R)-beta-phenylalanine with an enantiomeric purity of 90% e.e. was obtained. The initial step of the (R)-isomer degradation was supposed to be oxidative deamination. Resting cell reaction with vigorous shaking, with cells of Arthrobacter sp. as the catalyst, resulted in production of 0.49% (w/v) of (S)-beta-phenylalanine with an enantiomeric purity of 99% e.e. from 1.0% (w/v) racemic beta-phenylalanine in 45 h.     

A comparison of raw starch-digesting glucoamylase production in liquid and solid cultures of Rhizopus strains

Maximum growth for Rhizopus sp. A-11 was obtained at a zinc ion concentration of 0.7 ppm in a liquid medium. Glucoamylase (GA, EC 3.2.1.3) production in Rhizopus sp. A-11 was maximized at 710 U/ml, at the presence of 75 ppm for calcium and 0.7 ppm of zinc ions in liquid medium. Zinc ion is known as an essential biometal for Rhizopus growth; however, growth was inhibited by the zinc ion concentration, not maximized. Although calcium ion was not necessary to Rhizopus growth, GA production using Rhizopus sp. A-11 was markedly stimulated by calcium ion concentration over 75 ppm in the liquid medium. The GA productivity of the present liquid culture was about 4.4 times higher than that of the solid state culture, based on the unit starch amount in the liquid and solid media carbon source. The characteristics of the GA produced by the Rhizopus sp. A-11 liquid culture were interesting; that is, almost all the GA produced was classified as raw starch-digesting GA (GA-I). Secreted protein in the culture liquid after 30 h was nearly GA, and had a limited amount of impure protein. As a result, it was found that using a Rhizopus culture in a specified metal-ion regulated medium was an effective method for producing GA. Thus the present culture method was renamed the "metal-ion-regulated liquid culture method".     

Fungal degradation of aflatoxin B1

A number of fungal cultures were screened to select an organism suitable to be used in the detoxification of aflatoxin B1. They were co-cultured in Czapek-Dox-Casamino acid medium with aflatoxin B1 producing Aspergillus flavus. Several fungal cultures were found to prevent synthesis of aflatoxin B1 in liquid culture medium. Among these Phoma sp., Mucor sp., Trichoderma harzianum, Trichoderma sp. 639, Rhizopus sp. 663, Rhizopus sp. 710, Rhizopus sp. 668, Alternaria sp. and some strains belonging to the Sporotrichum group (ADA IV B14(a), ADA SF VI BF (9), strain 720) could inhibit aflatoxin synthesis by > or =90%. A few fungi, namely ADA IV B1, ADA F1, ADA F8, also belonging to the Sporotrichum group, were less efficient than the Phoma sp. The Cladosporium sp. and A. terreus sp. were by far the least efficient, registering <10% inhibition. The cultures which prevent aflatoxin biosynthesis are also capable of degrading the preformed toxin. Among these, Phoma sp. was the most efficient destroying about 99% of aflatoxin B1. The cell free extract of Phoma sp. destroyed nearly 50 microg aflatoxin B1 100 ml(-1) culture medium (90% of the added toxin), and this was more effective than its own culture filtrate over 5 days incubation at 28+/-2 degrees C. The degradation was gradual: 35% at 24 h, 58% at 48 h, 65% at 72 h, 85% at 96 h and 90% at 120 h. The possibility of a heat stable enzymatic activity in the cell free extract of Phoma is proposed.     

Production of flavour esters by Rhizopus oryzae

Microbial production of different alipathic esters with flavour characteristic has been studied. Lyophilized whole cells of Rhizopus oryzae CBS 112-07 were found to be particularly suitable to catalyse the synthesis of different flavour esters (hexyl acetate, propionate, butyrate, caprylate; geranyl acetate, propionate, butyrate and 2- and 3-methylbutyl acetate, butyrate) in n-heptane. This strain was therefore utilized for the semipreparative production of geranyl butyrate by semicontinous and continous addition of the substrates with satisfactory yields (144 g l^–1 in 264 h and 142 g l^–1 in 48 h respectively).     

Resolution of 2,2-diphenyl-1,3-dioxolane-4-methanol via Rhizopus sp. lipase-catalyzed enantioselective transesterification with vinyl butanoate

2,2-Diphenyl-1,3-dioxolane-4-methanol was resolved via Rhizopus sp. lipase-catalyzed enantioselective transesterification with vinyl butanoate in isopropyl ether (E=23). A gram-scale resolution resulted in the production of the (R)-alcohol with 95% e.e. in 33% yield based on racemate.     

黑根霉对人参皂苷Re的生物转化

利用菌种黑根霉Rhizopus sp.对人参皂苷Re进行生物转化,并对人参皂苷Re及其发酵产物进行HPLC系统分析比较,经液相色谱-质谱分析得出人参皂苷Re转化率为92.16%,并制备出人参皂苷Re发酵产物中峰值升高的成分,转化后的人参皂苷发酵产物中化合物1确定为人参皂苷Rg2,化合物2为Rg2的同分异构体,得率为10.13%;化合物3和化合物4确定为人参皂苷Rg5/Rk1,得率为29.23%。从结果初步推测得出人参皂苷Re被黑根霉转化为人参皂苷Rg2的机理,人参皂苷Re转化成人参皂苷Rg5/Rk1的机理还有待于进一步研究。     

Microbial production of optically active β-phenylalanine ethyl ester through stereoselective hydrolysis of racemic β-phenylalanine ethyl ester

The ability to produce (R)- or (S)-β-phenylalanine ethyl ester (3-amino-3-phenylpropionic acid ethyl ester, BPAE) from racemic BPAE through stereoselective hydrolysis was screened for in BPAE-assimilating microorganisms. Sphingobacterium sp. 238C5 and Arthrobacter sp. 219D2 were found to be potential catalysts for (R)- and (S)-BPAE production, respectively. On a 24-h reaction, with 2.5% (w/v) racemic BPAE (130 mM) as the substrate and wet cells of Sphingobacterium sp. 238C5 as the catalyst, 1.15% (w/v) (R)-BPAE (60 mM) with enantiomeric purity of 99% e.e. was obtained, the molar yield as to racemic BPAE being 46%. On a 48-h reaction, with 2.5% (w/v) racemic BPAE (130 mM) as the substrate and wet cells of Arthrobacter sp. 219D2 as the catalyst, 0.87% (w/v) (S)-BPAE (45 mM) with enantiomeric purity of 99% e.e. was obtained, the molar yield as to racemic BPAE being 35%. The enzyme stereoselectively hydrolyzing (S)-BPAE was purified to homogeneity from the cell-free extract of Sphingobacterium sp. 238C5. The enzyme was a monomeric protein with a molecular mass of about 42,000. The enzyme catalyzed hydrolysis of β-phenylalanine esters, while the common aliphatic and aromatic carboxylate esters were not catalyzed.