Synthesis of a Diastereomeric Mixture of 15, 19, 23-Trimethylheptatriacontane,the Most Active Component of the Sex Pheromone of the Female Tsetse Fly,Glossina morsitans morsitans

An α-amylase secreted by Pichia burtonii 15-1 isolated from a traditional starter murcha of Nepal, named Pichia burtonii α-amylase (PBA), was studied. The gene was cloned and its nucleotide sequence was determined. PBA was deduced to consist of 494 amino acid residues. It shared certain degrees of amino acid sequence identity with other homologous proteins: 60% with Schwanniomyces occidentalis α-amylase, 58% with Saccharomycopsis sp. α-amylase, and 47% with Taka-amylase A from Aspergillus oryzae. A three-dimensional structural model of PBA generated using the known three-dimensional structure of Taka-amylase A as a template suggested high structural similarity between them. Kinetic analysis revealed that the K_m values of PBA were lower than those of Taka-amylase A for the oligosaccharides. Although the k_cat values of PBA were lower than those of Taka-amylase A for the oligosaccharide substrates, the k_cat/K_m values of PBA were higher.     

Five New Phenolic Glycosides from Viburnum luzonicum

Viburnum luzonicum is widely distributed in China. Its branch extracts showed potential α-amylase and α-glucosidase inhibitory activities. In order to discover new bioactive constituents, five undescribed phenolic glycosides, viburozosides A−E ( 1 – 5 ), were obtained by bioassay-guided isolation coupled with HPLC-QTOF-MS/MS analysis. Their structures were elucidated by spectroscopic analyses, including 1D NMR, 2D NMR, ECD, and ORD. All compounds were tested for their α-amylase and α-glucosidase inhibitory potency. Compound 1 showed significantly competitive inhibition against α-amylase (IC₅₀=17.5 μM) and α-glucosidase (IC₅₀=13.6 μM).     

Pyrindicin,a New Alkaloid from a Streptomyces Strain

In the course of our examination for the alkaloid productivities of Streptomyces strains, Streptomyces sp. NA–15 was found to produce a new alkaloid, pyrindicin, in the culture medium. The strain NA–15 was found to be a variant of Streptomyces griseoflavus and was designated as S. griseoflavus var. pyr indie us nov. var.

After the culture conditions for pyrindicin production were studied, pyrindicin was obtained as its hydrochloride (mp 145°C, decomp.) from the cultured broth. The compound was shown to possess weak antimicrobial and several pharmacological activities. The LD₅₀ of the hydrochloride (ip, in mice) was 87 mg/kg.     

Purification,Crystallization and Properties of Tryptophanase from Proteus rettgeri

An inducible tryptophanase was crystallized from the cell extract of Proteus rettgeri grown in a medium containing l-tryptophan. The purification procedure included ammonium sulfate fractionation, heat treatment, DEAE-Sephadex and hydroxylapatite column chromatographies. Crystals were obtained from solutions of the purified enzyme by the addition of ammonium sulfate.

The crystalline enzyme preparation was homogeneous by the criteria of ultracentrifugation and zone electrophoresis. The molecular weight was determined to be approximately 210,000.

The crystalline enzyme catalyzed the degradation of l-tryptophan into indole, pyruvate and ammonia in the presence of added pyridoxal phosphate. The enzyme also catalyzed pyruvate formation from 5-hydroxy-l-tryptophan, 5-methyl-l-tryptophan, S-methyl-l-cysteine and l- cysteine. l-, d-Alanine, l-phenylalanine and indole inhibited pyruvate formation from these substrates.     

人胰腺α-淀粉酶抑制剂高通量筛选模型的建立及其应用

【目的】针对人胰腺α-淀粉酶这个糖代谢途径中重要的靶蛋白,建立α-淀粉酶抑制剂高通量筛选模型。【方法】采用毕赤酵母表达系统克隆和表达人胰腺α-淀粉酶;利用酶的催化特性建立α-淀粉酶抑制剂筛选模型;应用该模型对放线菌发酵液冻干物进行高通量筛选;通过构建16S rRNA系统发育树分析阳性菌株的分类地位。【结果】成功克隆、表达了具催化活性的人胰腺α-淀粉酶;建立了α-淀粉酶抑制剂的筛选模型;对近2000株放线菌的发酵液冻干物进行高通量筛选,最终得到14株α-淀粉酶抑制剂产生菌株,且在分类学上具有丰富的菌种多样性。【结论】本研究建立的α-淀粉酶抑制剂高通量筛选模型具有很强的实用价值,可用于新型淀粉酶抑制剂类降糖药物的开发。     

酶法提取玉米胚中总黄酮的研究

尝试使用α-淀粉酶进行糖苷转移提取玉米中总黄酮,同时对α-淀粉酶的固定化进行研究。对α-淀粉酶固定化的研究表明:以壳聚糖为载体,用体积分数4%戊二醛进行交联,加酶量为3 g.L-1条件下可以获得最佳的固定化效果;与游离酶相比,其最适作用温度范围、pH值范围均比游离酶范围宽;固定化酶的热稳定性优于游离酶,且具有良好的操作稳定性。本试验使用游离酶法、固定化酶法提取玉米中总黄酮,得率分别为2.82%、2.46%。     

Various solvent effects on phytochemical constituent profiles,analysis of antioxidant and antidiabetic activities of Hopea parviflora

The current research has been designed to assess the phytochemical composition, antioxidant and antidiabetic properties of Hopea parviflora, sequentially extracted with petroleum ether, chloroform, ethyl acetate, ethanol and methanol. All the five extracts were tested for qualitative and quantitative phytochemicals. DPPH, Superoxide, FRAP, ABTS and metal chelating antioxidant activities were evaluated. Antidiabetic potentials of all the five extracts were tested using standard in vitro α- amylase and α - glycosidase inhibition assays. Qualitative phytochemical screening showed the presence of alkaloids in all the extracts except petroleum ether and ethyl acetate. Steroids were present in the petroleum ether, ethyl acetate and chloroform extracts whereas glycosides were present in all the extracts, except ethanol. The total phenol, flavonoid, tannin and saponins contents varied from solvent to solvent, with the highest values being 18.9, 18.2, 0.98 and 39.9 mg/mL, respectively. Methanolic extract showed the highest antioxidant activities in DPPH, FRAP and superoxide assays. Moreover, effective results were observed for the ethanol and ethyl acetate extracts in the ABTS and metal chelating assays. The methanolic extract showed potential antidiabetic activities with the IC₅₀ values of 230.2 and 308.2 μg/mL in α- amylase and α -glycosidase inhibition assays, respectively.     

Benzaldehyde O-Alkyloximes as New Plant Growth Regulators

Fourty-two kinds of benzaldehyde O-alkyloximes derived from benzaldehydes were prepared and their biological activities were investigated. Introduction of a fluorine or bromine atom to the benzene ring of the oximes enhanced their phytotoxic activity. The O-alkyloximes with a fluorine atom at the 3 or 4 position of the benzene ring were more active than the other oximes in the GA₃-induced α-amylase induction inhibition test. In the transpiration test, 4-bromobenzaldehyde O-carboxylmethyloxime was the most active. The O-alkyloximes exhibited weak abscisic acid-like activity by inhibiting not only the germination, root growth and transpiration of higher plants but also GA₃-induced α-amylase induction in embryoless barley seeds.     

Fungicidal Activity of New Diphenylphosphinic Amide Derivatives

We isolated erythropoietin (Epo) from anemic-rat serum with 1.3 × 10⁶-fold purification and 38% recovery using immunoaffinity chromatography. The isolated Epo migrated in SDS polyacrylamide gel with a molecular size of 37 kDa. Biological properties of rat Epo were compared with those of human Epo using target cells of primate and murine origins. When murine cells were used as target cells for assaying Epo, rat Epo stimulated proliferation of the cells with a 50% lower potency than did human Epo. The activity of rat Epo on human cells was only 25% of that of human Epo. Studies of Epo binding to the receptor indicated that rat and human Epos were not distinguishable in binding to murine cells; however, rat Epo bound to the receptor on human cells with an affinity much lower than that of human Epo. Rat Epo was digested with N-glycanase. Complete removal of N-linked sugars converted the native Epo to the deglycosylated form with 18 kDa. The in vitro activity of deglycosylated Epo was 2.5-fold higher than that of the native Epo.     

Subsite Structure of α-Amylase II from Thermoactinomyces vulgaris R-47

We estimated the subsite structure of α-amylase II (TVA II) from Thermoactinomyces vulgaris R-47 expressed in Escherichia coli. TVA II has eight subsites, and the catalytic site is between the 5th and 6th subsite from the non-reducing end side. The subsite affinities, A_-5, A_-4, A_-3, A_-2, (A_-1+A₊₁), A₊₂, and A₊₃, were calculated to be -0.35, 0.93, 0.55, 2.56, 1.18, 1.71, and 0.01 kcal mol^-1, respectively.     

Diphaladine A，扁枝石松中一个新的石松生物碱

从扁枝石松（Diphasiastrum complanatum）的95％乙醇提取物中分离得到1个新的石松生物碱,经波谱技术鉴定其结构,命名为diphaladine A（1）。同时还分离得到其它9个已知石松生物碱类化合物,其中obsctmmaine A（2）,L20,lycoposerramine—K,des—N—methyl—β—obscurine,des-N—methyl-α—obscurine,lycoflexion,和phlegmariurine B等7个化合物均为首次从该植物中分离得到。     

Chaperone action of a versatile small heat shock protein from Methanococcoides burtonii, a cold adapted archaeon

The Methanococcoides burtonii small heat shock protein (Mb-sHsp) is an alphaB-crystallin homolog that delivers protein stabilizing and protective functions to model enzymes, presumably reflecting its role as a molecular chaperone in vivo. Although the gene encoding Mb-shsp was cloned from a cold-adapted microorganism, the Mb-sHsp is an efficient protein chaperone at temperatures far above the optimum growth temperature of M. burtonii. We show that Mb-sHsp can prevent aggregation in E. coli cell free extracts at 60 degrees C for 4 h and can stabilize bovine liver glutamate dehydrogenase for 3 h at 50 degrees C. Surface plasmon resonance was used to determine the binding affinity of Mb-sHsp for denatured proteins. Mb-sHsp bound tightly to denatured lysozyme but not to the native form. When Mb-Cpn and Mg(2+)-ATP were added to the reaction, bound lysozyme was released from Mb-sHsp establishing that Mb-Cpn is able to off-load folding intermediates from Mb-sHsp. In addition, Mb-sHsp and Mb-Cpn also function cooperatively to protect an enzyme substrate. Through characterization of these M. burtonii chaperones, we were able to reconstitute a key heat shock regulated protein folding function of this cold adapted organism in vitro.     

Polymorphism in Rice Amylases at an Early Stage of Seed Germination

A polymorphism in rice amylases at an early stage of seed germination is analyzed by zymogram. In nonglutinous cultivars of rice, α-amylase isozymes are mainly confirmed in germinating seeds. However, in glutinous cultivars, β-amylase isozymes, which are not confirmed in nonglutinous cultivars, make up the major part of the total amylase activity and the expression of α-amylases are repressed.     

Identification of a coffee berry borer‐associated yeast: does it break down caffeine?

Two yeasts isolated from laboratory reared adult coffee berry borers [Hypothenemus hampei (Ferrari) (Coleoptera: Scolytidae)] and from insects collected in the field in Colombia were identified as Pichia burtonii Boidin and Candida fermentati (Saito) Bai, based on sequencing of the nuclear large subunit 26S rDNA variable D1/D2 domain. Liquid culture experiments using P. burtonii in media containing different caffeine levels indicated that caffeine levels in a range found within coffee seeds can retard yeast growth. HPLC analysis shows that P. burtonii does not break down caffeine.     

Continuous-flow determination of an alpha-amylase inactivator in fermentation samples using a ferricyanide reagent

A continuous-flow system has been developed in which pancreatic alpha-amylase is incubated with soluble starch at 37 degrees C. Reducing sugars being delivered at the "steady-state" hydrolysis of starch are dialyzed into a solution of alkaline ferricyanide. Ferricyanide is then reduced at 95 degrees C. The decrease in the absorbance of ferricyanide solution is recorded as "enzyme baseline." When samples containing alpha-amylase inactivator are introduced into the system, hydrolysis of starch is reduced according to the concentration of enzyme inactivator. The amount of inactivator in the samples is deduced from a standard curve of six standard concentrations using the Technicon Autoanalyzer II system manager in connection with a calculator for automated interpolation. This method allows to determine 40 samples/h with high sensitivity and precision (mean = 1.53 mg/liter; C.V. = 1.4%).     

The effect of GNA lectin on the α-amylase activity of the beet armyworm,Spodoptera exigua Hb. (Lepidoptera: Noctuidae)

Beet armyworm (Spodoptera exigua Hb.) (Lepidoptera: Noctuidae) is the major pest of sugar beet (Beta vulgaris). Pesticide applications are the main method of the insect control. So, alternative method/s is/are needed to control this insect species. So, in the current study, the effect of Galanthus nivalis agglutinin (GNA) (snowdrop lectin) on beet armyworm α-amylase was studied. Measurement of the amylase activity of the larval midgut fed on artificial diet and sugar beet leaves showed that the enzyme activity was higher when the larvae fed on artificial diet. However, in both cases, the fourth instar larvae had the greatest amylase activity. Thus, fourth instar larvae were offered artificial diet containing 1 and 2% GNA. Both treatments of the lectin significantly reduced the α-amylase activity of the insect. For example, amylase activity of the fourth instar larvae in the control (fed only on artificial diet) was 2.62 Uml^?1 whilst the activity of the enzyme in the two treatments including diet containing 2 and 1% lectin was 1.45 and 1.75 Uml^?1, respectively. The achieved data showed that lectin, in addition to have toxic effect on the larval growth and development, affects the α-amylase activity of the insect gut.     

东川乌头中一个新的去甲二萜生物碱( 英文)

从东川乌头 (AconitumgeniculatumFletcheretLauener)块根的乙醇提取物中分离得到 3个去甲二萜生物碱 ,经 1D、 2D -NMR技术鉴定 ,分别为 2 0 -乙基 - 8-乙酰氧基 - 14- (对 -羟基苯甲酰氧基 ) - 1α ,6α ,16 β ,18-四甲氧基乌头烷 - 3α ,13β二醇 (1)、 2 0 -乙基 - 8-乙酰氧基 - 14-苯酯基乌头烷 - 3α ,13β二醇 (2 )和 2 0 -乙基 - 8-乙酰氧基 - 14- (对 -甲氧基苯酯基 )乌头烷 - 3α ,13β二醇 (3) ,其中 1为新化合物 ,命名为滇羟碱 (geniculine)。     

A New Diterpenoid Alkaloid from Aconitum leucostomum

Four diterpenoid alkaloids were isolated by means of column chromatography and preparative HPLC from the roots of Aconitum leucostomum Worosch. Their structures were determined from the spectral data. One of them was found to be a new compound named leucostine ( Ⅱ ). The other three are known alkaloids, leuconine ( Ⅰ ), dehydroacosanine ( Ⅲ ) and sepaconitine (IV).     

Pentane Inhibition of the Egg-white Lysozyme-induced Cell Lysis of Micrococcus lysodeikticus

Pentane inhibited the cell lysis of Micrococcus lysodeikticus by egg-white lysozyme, when added to the lysozyme solution before mixing the cells. The pentane inhibition was not observed when pentane was added either before to the cell suspensions or to the cell-lysozyme mixture. The degree of pentane inhibition was proportional to the concentration of pentane, and the maximum inhibition was achieved with about 5% pentane close to the saturation point. On the other hand, pentane did not inhibit the hydrolysis of glycol chitin by the lysozyme, showing that the β-1,4-glucosaminidase activity of lysozyme remained unchanged. The inhibitory action of pentane on the lysozyme-induced cell lysis was of a competitive nature. The pentane inhibition had no ph-dependence, but it was influenced by the ionic strength of the buffer used as solvent.

When the lysozyme solution was treated with pentane, a characteristic ultraviolet spectrum of lysozyme was produced; a blue shift with a minimum at 280 nm and a trough at 291 nm. The degree of spectral change at 280 nm depended on the concentration of pentane. Pentane was, therefore, found to interact with egg-white lysozyme to affect the conformation and enzymatic activity of lysozyme. The mechanism of pentane inhibition on the lysozyme-induced cell lysis was discussed.     

软珊瑚Nephthea sp.中新生物碱分离与结构鉴定(英文)

一个新的化合物Nephoxaloid(1)具有独特的三环骨架结构和一个已知的化合物蕨藻素Caulerpin(2)从中国国西沙群岛的软珊瑚Nephthea sp.中分离得到。通过NMR和MS对它们的平面结构进行分析,化合物(1)和(2)对少数癌细胞株具有细胞毒性作用也有报道。