Inhibition of Candida rugosa lipase by saponins, flavonoids and alkaloids

Lipase inhibitors have generated a great interest because they could help in the prevention or the therapy of lipase-related diseases. Therefore, the aim of the work was to evaluate by HPLC, and using Candida rugosa lipase as model, the inhibitory effect of several saponins: β-aescin, digitonin, glycyrrhizic acid (GA) and Quillaja saponin (QS); flavonoids: 3-hydroxyflavone, 5-hydroxyflavone, (±)-catechin and kaempferol; and alkaloids: aspidospermine, papaverine, physostigmine, pilocarpine, raubasine, rescinnamine, reserpine and trigonelline.

The inhibition produced by most of these compounds is described here for the first time. Saponins appeared very active, being β-aescin and digitonin the most active compounds (IC₅₀ = 0.8–2.4 × 10⁻⁵ M). The inhibitory activity of flavonoids was lower than that of saponins (except GA), and (±)-catechin and kaempferol were the most active. Alkaloids was the most heterogeneous group assayed, varying from rescinnamine, with an IC₁₆ similar to that of digitonin, to papaverine and others which showed almost no inhibition.

In conclusion, β-aescin, digitonin, kaempferol or (±)-catechin, strong lipase inhibitors with a low toxicity and present herbal drugs used for lipase-related diseases such as acne or ulcer, are promising candidates for the prevention or the treatment of these diseases.     

Lipase inhibitors for the treatment of acne

Propionibacterium acnes lipase is an important factor in the pathogenesis of acne, because free fatty acids formed as a result of the effect of P. acnes lipase on sebaceous triglycerides induce severe inflammation. Tetracyclines have been the most common systemic therapy for acne due to their beneficial clinical effects, their inhibition of lipase activity of P. acnes as well as their inhibition of P. acnes chemotaxis. Erythromycin and Jumi-haidoku-to also inhibit the lipase activity of P. acnes. Among P. acnes biotypes and serotypes, stronger P. acnes lipase activities were seen in P. acnes biotype III and serotype II. Conversely, Propionibacterium granulosum is also isolated in acne lesions but has only a faint lipase activity, which might not be of significance. In the future, the degree of P. acnes lipase activity in acne, identified by genetic techniques, needs to be compared to the condition of acne rash.     

珠子参中法尼基焦磷酸合酶(FPS)对皂苷生物合成的影响研究

珠子参作为名贵中草药已有上千年应用历史,珠子参皂苷是珠子参的主要活性成分,由达玛烷型和齐墩果烷型三萜皂苷组成。目前,对珠子参皂苷的生物合成了解甚少,有必要开展与皂苷合成相关的基础研究工作。已有报道表明,法尼基焦磷酸合酶（FPS）可能是植物中皂苷合成的关键酶。本研究克隆了珠子参FPS基因（PjFPS）的cDNA序列,并对其进行了生物信息学分析。基于PjFPS序列构建了珠子参过表达载体pCAMBIA2300s-PjFPS,将其转化到珠子参细胞中,成功获得阳性转基因细胞;测定了阳性细胞系中PjFPS基因的相对表达量、FPS酶活、皂苷以及植物甾醇含量的变化。结果表明,与野生型珠子参细胞相比,PjFPS转基因珠子参细胞系中,PjFPS基因的相对表达量、FPS酶活以及皂苷含量均有不同程度的提高。而且,由于皂苷合成代谢流的增加,也促进了相关重要关键酶基因的表达。在效果最好的阳性细胞中,PjFPS基因的相对表达量、FPS酶活、皂苷含量分别为对照的12、4和3倍;同时,转基因细胞系的植物甾醇含量也有所升高。本研究通过对皂苷合成途径关键酶基因的调控实现了对皂苷生物合成的调节,为获得高效、稳定的珠子参皂苷合成调控技术提供理论参考和依据。     

Prey-Dependent Life Attributes of an Aphidophagous Ladybird Beetle, Propylea dissecta (Coleoptera: Coccinellidae)

Predation potential, development, immature survival and reproduction of an aphidophagous ladybeetle, Propylea dissecta (Mulsant) was studied when fed on seven aphid prey, viz. Aphis gossypii, Aphis craccivora, Lipaphis erysimi, Uroleucon compositae, Brevicoryne brassicae, Rhopalosiphum maidis and Myzus persicae. A. gossypii was most suitable and consumed by the larvae and adults of P. dissecta, while M. persicae, the least. Pre-imaginal development of P. dissecta was fastest (0.080 day^-1) when A. gossypii was used as prey, whilst slowest (0.061 day^-1) on M. persicae. The immature survival, adult emergence, adult male and female longevity of P. dissecta was maximal (i.e., 77.10±0.04 and 93.21±0.79%, 57.10±1.62 and 62.40±1.93 days, respectively) on A. gossypii and minimal (i.e., 63.01±1.87 and 81.73±1.79%, 42.50±1.21 and 49.40±2.32 days, respectively) when M. persicae was provided as prey. Oviposition period, fecundity, percent egg viability and mean reproductive rate was maximum (i.e., 50.30±2.03 days, 856.00±30.00 eggs, 96.40±0.31% and 17.02 eggs per day) on A. gossypii, and minimum (i.e., 18.00±1.40 days, 212.00±18.21 eggs, 72.46±2.81% and 11.78 eggs per day) on M. persicae. Adult weight and developmental rate of P. dissecta have a positive correlation, which suggests that if immature stages of ladybeetle developed faster, they should grow into heavier adults. Female longevity and fecundity also have a positive correlation. The findings also reveal that all seven aphid species tested are essential food. Rank order of prey species was consistent in all experimental parameters.     

In vitro antiplasmodial activity of extract and constituents from Esenbeckia febrifuga, a plant traditionally used to treat malaria in the Brazilian Amazon

Esenbeckia febrifuga (Rutaceae) is a plant traditionally used to treat malaria in the Brazilian Amazon region. Ethanol extract of stems displayed a good antiplasmodial activity against Plasmodium falciparum strains W-2 (IC₅₀ 15.5±0.71 μg/ml) and 3 D7 (IC₅₀ 21.0±1.4 μg/ml). Two coumarins (bergaptene 1 and isopimpinellin 2), five alkaloids (flindersiamine 3, kokusaginine 4, skimmiamine 5, γ-fagarine 6 and 1-hydroxy-3-methoxy-N-methylacridone, 7), besides a limonoid (rutaevine 8), have been isolated for the first time from this species. Antiplasmodial activity of compounds 3, 5–8 has been evaluated in vitro against P. falciparum strains (W-2 and 3D7) and the furoquinolines 5 and 6 were the most potent displaying IC₅₀ values <50 μg/ml; flindersiamine (3) showed a weak activity while alkaloid 7 and rutaevine (8) were inactive (IC₅₀>100 μg/ml).     

Characterization of 1,3-regiospecific lipases from new Pseudomonas and Bacillus isolates

Lipase producing ability of 120 bacterial isolates was examined qualitatively, resulting in 32 lipase producers, which were further screened for 1,3-regiospecificity. Three Bacillus (GK-8, GK-31 and GK-42) and one Pseudomonas (GK-80) were found to produce 1,3-regiospecific lipases. These lipases were alkaline in nature as they showed pH optima of 9.0 and high stability in the alkaline pH range of 8.0–11.0. The lipases from three Bacillus isolates, viz. GK-8, GK-31 and GK-42 showed temperature optima of 37 °C, whereas the Pseudomonas (GK-80) lipase showed optimum activity at 50 °C. The lipase of GK-8 was highly stable and showed enhanced activity in different organic solvents like petroleum ether (172%), diethyl ether (143%) and acetone (135%).     

Effects of culture media, temperature, pH, and light on growth, sporulation, germination, and bioherbicidal efficacy of Phoma exigua, a potential biological control agent for salal (Gaultheria shallon)

In order to evaluate the potential use of Phoma exigua isolate PFC 2705 (PFC2705) as a biological control agent for salal (Gaultheria shallon), effect of cultural and environmental parameters on growth, conidia production, and pathogenicity of P. exigua were characterized in studies conducted under laboratory and greenhouse conditions. Within a range of 5-30°C, the optimum growth and germination temperature range was 20-25°C. The effect of pH on mycelial growth and conidial germination was not significant from pH 5 to 10. Fluorescent light significantly enhanced sporulation of the fungus on most agar media tested, yet was not necessary for growth. The type of culture media significantly affected mycelium growth, sporulation, and conidia germination. Age of mycelia used as inoculum affected the disease severity on salal. PFC2705 suppressed the growth of mature salal plant by inciting lesions on leaves, branch tips, and axillary buds and caused 56% death of the total biomass above ground. Characteristics such as easy inoculum production, wide range of growth environments, and high infectivity on salal increased the potential of P. exigua as a biocontrol agent for management of salal.     

Anti Propionibacterium acnes activity of rhodomyrtone, an effective compound from Rhodomyrtus tomentosa (Aiton) Hassk. leaves

Propionibacterium acnes have been recognized as one of the main causative agents in pathogenesis of acne. Twenty one isolates of P. acnes isolated from acne lesions were screened for lipase and protease activity which are reported to be associated in acne and inflammation. Interestingly, all P. acnes isolates demonstrated lipase activity. Similarly, 90% of test P. acnes produced protease enzyme. Antibacterial activity of the ethanol extract of Rhodomyrtus tomentosa (Aiton) Hassk. leaves and rhodomyrtone, its principle compound were tested against P. acnes using broth macrodilution method. The MIC(90) values of the ethanol extract and rhodomyrtone were 32 and 0.5 μg/mL, respectively. The numbers of the bacterial cells were reduced at least 99% after treatment with the ethanol extract and rhodomyrtone within 72 and 24 h, respectively. Cytotoxicity test of the extract and rhodomyrtone was performed on human normal fibroblast. The IC(50) values of the ethanol extract and rhodomyrtone were 476 and more than 200 μg/mL, approximately 15 and 400 folds higher than the MIC(90) values indicating that both substances were very low cytotoxic which could be applied as topical therapeutic anti-acne agents.     

Influence of oxygen on ethanol and xylitol production by xylose fermenting yeasts

The behaviour of Pichia stipitis, Pachysolen tannophilus, Candida shehatae and Candida parapsilosis was investigated to select the most suitable yeast to convert xylose either to ethanol or to xylitol, with little or no formation of by-products. The aeration rate was used as a variable parameter. P. stipitis and C. parapsilosis were the most effective producers or ethanol and xylitol, respectively, both reaching productivities at very low levels of oxygenation. With P. stipitis, better ethanol productivity was attained under microaerobic conditions (K_La = 4·8 h⁻¹) while with C. parapsilosis high yields and rates of xylitol production were detected at K_La values of about 16·3 h⁻¹. P. tannophilus and C. shehatae showed lower performances under all conditions used while changes in oxygenation modified the ratio of ethanol to xylitol produced by these yeasts, suggesting that they are more dependent on the oxygen power input than P. stipitis and C. parapsilosis. The influence of oxygen transfer rates on ethanol and xylitol formation with the best producers is discussed.     

3种速生树种落叶水浸提液对走马胎幼苗生长的化感效应

林药复合种植的关键在于科学理解造林树种与药材之间是否具有增益效应,其中避免化感抑制是重要方面之一。本研究选择以民族药用植物—走马胎（Ardisia gigantifolia Stapf）为研究对象,探讨走马胎与3种速生树种尾叶桉（Eualyptus urophylla S.T.Blakely）、湿地松（Pinus elliottii Engelm.）和杉木（Cunninghamia lanceolata（Lamb.）Hook.）复合种植中存在的化感作用。结果显示：与对照组CK相比,桉树落叶水浸提液降低了走马胎株高至少2.59%,对走马胎幼苗的株高有显著性抑制作用（P<0.05）,对走马胎幼苗的根长、地下部分干重以及生物量有极显著性抑制作用（P<0.01）,且随着浓度的增高,抑制作用增强。湿地松落叶水浸提液对走马胎幼苗的株高、根长以及地上部分干重出现低浓度促进高浓度没有显著性影响的双浓度效应,而对走马胎幼苗的基径、地下部分干重以及生物量呈现低浓度促进高浓度抑制的双浓度效应;杉木落叶水浸提液对走马胎幼苗生长的影响呈现低浓度促进高浓度抑制的双浓度效应现象。综合各项指标得出：走马胎与桉树之间存在较强的负化感效应,而走马胎与湿地松或杉木之间存在“低促高抑”的化感效应;     

Structured fiber supports for gas phase biocatalysis

Pseudomonas cepaciae lipase adsorbed onto non-porous structured fiber supports in the form of woven fabrics, was used to catalyze hydrolysis and transesterification reactions in the gas phase. The enzyme adsorbed onto carbon fiber support exhibited much higher catalytic activity compared to the enzyme immobilized onto glass fiber carrier. The effect of temperature and relative humidity on reactions catalyzed by P. cepaciae lipase adsorbed onto structured fiber carbon support was studied in the gas system. Under the conditions investigated (up to 60 °C and 80% relative humidity), the immobilized enzyme showed a high thermostability and could be efficiently used to catalyze hydrolytic and transesterification reactions in continuous mode. Structured fiber supports, with a high specific surface area and a high mechanical resistance, showed a low-pressure drop during the passage of reactants through a reactor. The approach proposed in this study could be suitable for immobilization of a wide variety of enzymes.     

Flora of the forest-steppe zone of West Siberia as a source of biologically active compounds

Plants of 76 species from 25 families growing in the forest-steppe zone of West Siberia are analyzed for the content of low-molecular biologically active compounds, such as flavonoids, tannins, catechins, coumarins, saponins, and alkaloids. It is established that 33 species hold promise as a source of flavonoids, 29 species tannins, 21 coumarins, 13 saponins, 13 alkaloids, and 1 catechins.     

Antimalarial properties of soy-bean fat emulsions

Intralipid^® and Ivelip^® are commercial preparations of soy-bean lipid extracts used for intravenous supplementation of lipids in various clinical conditions. They were found to inhibit the growth of Plasmodium falciparum in culture with an IC₅₀ of 8.07 ± 2.13 and 13.32 ± 2.05 mg.ml⁻¹, respectively. Intralipid^® rapidly and efficiently inhibited nucleic acid synthesis in cultured P. falciparum, exhibiting full inhibitory activity in less than 2 h. Ivelip^® injected intraperitoneally, was found by the 4-day suppressive test to be active in vivo against P. vinckei petteri within the normal recommended regimen for dietary lipid supply (0.5–4 g.kg⁻¹), but it was impossible to obtain a radical cure even with very high doses (6.4 g.kg⁻¹). Ivelip^® was less effective against P. berghei and P. yoelii nigeriensis. As Ivelip^® showed no interference with the antimalarial activity of chloroquine, it could be considered for use in the treatment of severe human malaria in association with 4-aminoquinolines to expedite the clearance of parasites.     

Biology of Coeloides sordidator (Hymenoptera: Braconidae), a Possible Candidate for Introduction Against Pissodes strobi (Coleoptera: Curculionidae) in North America

Various aspects of the biology of Coeloides sordidator, an ectoparasitoid of pine weevils of the genus Pissodes, were studied. When reared in the laboratory on P. castaneus, females preferred to oviposit on third and fourth instar larvae in galleries, but ovipositions were also observed on second instars and prepupae. Pupae were not accepted as hosts. At 23°C and a 16 h day length, pre-imaginal development lasted 15-20 days. For each parasitoid population tested, between 8 and 28% of the individuals diapaused as prepupae in cocoons. The rate of diapause was not influenced by the photoperiod during larval development. In contrast, diapause was induced by a short day photoperiod on parents before and during oviposition and by low temperatures during the larval stage. Diapause termination was achieved by cold storage at 2°C for at least 8 weeks. Rearing of C. sordidator on P. castaneus under natural conditions showed that in the Swiss Jura it is mainly univoltine, with less than 10% of the parasitoids emerging for a second generation. When C. sordidator females were offered healthy P. castaneus larvae and larvae parasitized by the braconid endoparasitoid Eubazus semirugosus, there was no evidence that they were able to discriminate between healthy and parasitized hosts. The potential of C. sordidator as a biological control agent against the North American species P. strobi is discussed.     

Geovanine,a new azaanthracene alkaloid from Annona ambotay aubl.

The new azaanthracene geovanine was isolated from the trunkwood of Annona ambotay Aubl. together with liriodenine, O-methylmoschatoline and other known substances, the flavonoids kaempferol, quercetin, (+)-dihydrokaempferol, (+)-dihydroquercetin, (±)-eriodictiol and (+)-catechin and the steroids sitosterol and 5α-stigmastan-3,6-dione.     

Diurnal variations in physiology and behaviour of the noble crayfish Astacus astacus and the signal crayfish Pacifastacus leniusculus

Heart rate, locomotor activity, and oxygen consumption were recorded simultaneously and continuously in seven individuals of the noble crayfish Astacus astacus (Linneus 1758) and seven individuals of the signal crayfish Pacifastacus leniusculus (Dana 1852). The recordings were made in the laboratory over 7 days at 15°C under a 12 : 12 h dark : light regime. Circadian rhythms in heart rate, locomotor activity and oxygen consumption were found both in A. astacus and P. leniusculus. Increased heart rate, locomotor activity, and oxygen consumption levels during night time in both A. astacus and P. leniusculus illustrated expression of nocturnal behaviour. No differences in oxygen consumption levels were observed between A. astacus and P. leniusculus. Also, no significant difference between heart rate levels or heart rate variances was found in A. astacus and P. leniusculus at night. During day, however, heart rate levels, heart rate variances and locomotor activity were higher in P. leniusculus than in A. astacus. The higher activity level in P. leniusculus than in A. astacus during daytime indicates that P. leniusculus is less strictly nocturnal than is A. astacus.     

Pediocin SA-1, an antimicrobial peptide from Pediococcus acidilactici NRRL B5627: production conditions, purification and characterization

Fermentation broths of Pediococcus acidilactici NRRL B5627 exhibited a certain antimicrobial activity due to a bacteriocin produced during early growth and until the stationary phase of growth was reached (at optimum of 60% dissolved oxygen saturation). Its size was determined by electrospray ionization mass spectrometric analysis as 3.660 kDa. N-terminal sequencing showed that the bacteriocin had 19 amino acid residues in the order KYYGXNGVXTXGKHSXVDX. The purified bacteriocin is similar to pediocins isolated by various Pediococci and therefore, it belongs to the class IIa of bacteriocins and is thus designated pediocin SA-1. Sensitivity of the purified pediocin to various storage temperatures and enzyme treatments was examined. Purified pediocin SA-1 is heat stable for up to 60 min at 121 °C. Pediocin SA-1 is inhibitory to several food-borne pathogens and food spoilage bacteria. It appears to be significantly more effective against Listeria spp. compared to pediocin PD-1 produced by P. damnosus. The mode of action of the purified bacteriocin appears to be bactericidal.     

An inducible release of reactive oxygen radicals in four species of gorgonian corals

The capability for physical injury or heat stress to elicit the production of reactive oxygen species was examined in four species of gorgonian corals. The sea plumes Pseudopterogorgia elisabethae, Pseudopterogorgia americana, the sea rod Eunicea fusca and the azooxanthellate red branching gorgonian Lophogorgia chilensis were physically injured using sonic sound cavitations and heat shocked by incubation in 33°C sea water. The pharmacological probe, diphenylene iodonium chloride (DPI), an inhibitor of NAD(P)H oxidase and peroxidases was used to identify an enzymatic surrogate of the oxidative burst. Both injury and heat stress were capable of inducing the release of reactive oxygen species (ROS) in all gorgonians tested, yet the kinetics and amplitude of ROS release varied among genera. In both the treatments, P. americana demonstrated the largest oxidative burst among the other corals tested.     

红松查尔酮合成酶基因CHS密码子偏好性分析

查尔酮合成酶(Chalcone synthase,CHS)广泛存在于植物体内,是花色素形成过程中一种重要的酶,可以进一步催化生成黄酮类化合物。本研究采用Codon W和EMBOSS在线软件对红松查尔酮合成酶基因CHS的密码子使用偏好性进行分析,并与北美乔松等其他24种植物的CHS基因以及模式植物基因组进行比较,对认识红松CHS基因的密码子使用偏好性,为选择适宜的表达系统奠定了一定的基础。研究结果表明:红松CHS基因编码区的有效密码子数(ENC)和GC含量分别为48.92和0.548,C+G含量高于A+T含量,密码子偏好以A/T结尾;多数植物CHS基因的G+C含量高于A+T含量,且密码子更偏好C/G结尾;聚类分析表明,红松与马尾松和赤松的密码子使用偏好性的相似性较高;密码子使用频率研究发现,红松CHS遗传转化与异源表达较优的受体可能是大肠杆菌和拟南芥。     

毕赤酵母底盘芳香族氨基酸合成途径改造生产肉桂酸及对香豆酸*

目的:改造毕赤酵母使其异源合成类黄酮生物合成途径的重要中间体肉桂酸、对香豆酸,并优化前体芳香族氨基酸生物合成途径以提高毕赤酵母的生产能力。方法:在毕赤酵母GS115中利用乙醇诱导型人工转录系统表达Rhodotorula glutinis来源的苯丙氨酸解氨酶,并在该重组菌株中分别过表达胞内芳香族氨基酸生物合成途径中的关键酶或其突变体以进行优化。结果:异源表达苯丙氨酸解氨酶可使毕赤酵母将自身产生的L-苯丙氨酸、L-酪氨酸转化为肉桂酸(38.8 mg/L)、对香豆酸(34.2 mg/L),而通过过表达相关酶进行优化,最终肉桂酸和对香豆酸的产量分别达到124.1 mg/L和302.0 mg/L。结论:利用新的异源宿主毕赤酵母成功合成了肉桂酸、对香豆酸,并对胞内的芳香族氨基酸生物合成途径进行了优化,表明毕赤酵母具有生产黄酮类化合物的应用潜力,也为其他芳香族氨基酸衍生物或植物化合物在毕赤酵母中的异源合成奠定了基础。