The effect of α-melanocyte stimulating hormone on endotoxin-induced intestinal injury

We investigated the effect of α-melanocyte stimulating hormone (α-MSH) on endotoxin-induced intestinal inflammation and the role of nitric oxide and prostaglandins in this response. α-MSH treatment (25 μg/rat, intraperitoneally (i.p.); twice daily) reduced the severity of the lesions macroscopically and microscopically. This protective effect was found to be confined mainly to the distal ileum. These lesions were reversed by pretreatment with the non-selective COX inhibitor indomethacin (10 mg/kg, subcutaneously (s.c.)) but not by the selective COX-2 inhibitor nimesulide (3 mg/kg, s.c.), the NO donor sodium nitroprusside (4 mg/kg, i.v.) or the iNOS inhibitor dexamethasone (3 mg./kg, i.p.) at macroscopic level and reversed by Indo or Dex at microscopic level. Increased peroxidase activity -index of tissue neutrophil infiltration- in the distal ileum of LPS-treated rats was decreased by α-MSH and this effect was reversed by pretreatment with Indo. In conclusion, the neuropeptide α-MSH has a beneficial effect on endotoxin-induced distal intestinal lesions by a mechanism which probably involves nitric oxide and COX-1 derived prostaglandins.     

Conversion of α-methyltropate to optically active α-phenylpropionate by tropate-degrading Rhodococcus sp. KU1314

Microorganisms which can assimilate tropate were screened from soil. Among them, we found a microorganism which has an ability to convert α-methyltropate to optically active α-phenylpropionate, and it was identified as Rhodococcus sp. KU1314. Substrate specificity of the microorganism has been studied. When the aryl group was phenyl, 4-methoxyphenyl and 2-naphthyl, the substrate gave optically active α-propionate in good yields. To estimate the reaction mechanism, some compounds considered to be the intermediates were subjected to the reaction. Both enantiomers of α-methyltropate were converted to (R)-α-phenylpropionate with almost the same enantiomeric excess (68 and 72% from R-and S-enantiomers, respectively) and yield (605 and 48% from R-and S-enantiomers, respectively).     

In vitro antimicrobial activity of alpha-melanocyte stimulating hormone against major human pathogen Staphylococcus aureus

Alpha-melanocyte stimulating hormone (α-MSH) is an endogenous anti-inflammatory peptide reported to possess antimicrobial properties, however their role as antibacterial peptides is yet to be established. In the present study, we examined in vitro antibacterial activity of α-MSH against S. aureus strain ISP479C and several methicillin-sensitive (MSSA) and methicillin-resistant (MRSA) S. aureus strains. Antibacterial activity was examined by varying several parameters, viz., bacterial cell densities, growth phase, pH, salt concentration, and temperature. Antibacterial activity was also examined in complex biomatrices of rat whole blood, plasma and serum as well as in biofilm form of bacteria. Our results showed that α-MSH possessed significant and rapid antibacterial activity against all the studied strains including MRSA (84% strains were killed on exposure to 12 μM of α-MSH for 2 h). pH change from 7.4 to 4 increased α-MSH staphylocidal activity against ISP479C by 21%. Antibacterial activity of α-MSH was dependent on bacterial cell density and independent of growth phase. Moreover, antimicrobial activity was retained when α-MSH was placed into whole blood, plasma, and serum. Most importantly, α-MSH exhibited antibacterial activity against staphylococcal biofilms. Multiple membrane permeabilization assays suggested that membrane damage was, at least in part, a major mechanism of staphylocidal activity of α-MSH. Collectively the above findings suggest that α-MSH could be a promising candidate of a novel class of antimicrobial agents.     

Kinetics of the diacetyl and 2,3-pentanedione reduction by diacetyl reductase (α-diketone reductase (NAD)) from Staphylococcus aureus

The kinetic mechanism of diacetyl and 2,3-pentanedione reduction by diacetyl reductase from Staphylococcus aureus was investigated. The shape of the primary double reciprocal plots, the product inhibition pattern, and the features of the inhibition by a substrate analogue (acetone) show that diacetyl is reduced via an Ordered Bi-Bi mechanism, and 2,3-pentanedione by an Ordered Bi-Bi or Theorell-Chance mechanism. NADH is the leading substrate in both reactions.Affinity constants for the coenzyme and the substrates and inhibition constants for NAD, acetoin, and acetone were also calculated. This enzyme has a high affinity for NADH; K_m (31–50 μM) and K_s (20–27 μM) for this compound are around one-tenth of the NADH intracellular concentration. Therefore, it must operate in vivo saturated with the coenzyme. This condition is not adequate to play the role, formerly proposed for diacetyl reductases, of regulating the equilibrium between oxidized and reduced forms of pyridine-nucleotides.     

金黄色葡萄球菌肠毒素中毒的免疫治疗策略

金黄色葡萄球菌肠毒素（staphylococcal enterotoxins,SEs）是由金黄色葡萄球菌（Staphylococcus aureus）分泌的一类具有呕吐活性的细菌外毒素，可引起严重的炎症反应和食物中毒。SEs具有超抗原活性，可与T细胞受体的可变区和MHC II类分子形成三元复合物（TCR-SEs-MHC II），直接刺激T淋巴细胞大量产生肿瘤坏死因子-α (tumor necrosis factor α,TNF-α)、白介素（interleukin,IL）-2、IL-6和γ干扰素（interferon γ,IFN-γ）等细胞因子，从而导致中毒性休克综合征（toxicshocksyndrome, TSS）。在临床常见的SEs中，肠毒素A (staphylococcalenterotoxin A, SEA)和肠毒素B(staphylococcal enterotoxin B,SEB)是出现频率最高、毒性最大、危害最严重的两种。目前尚没有针对SEs中毒治疗策略的综述性研究。本文首先概述了SEs的分类、结构及毒性作用，重点围绕SEA和SEB，分析了TCR-SEs-MHC II类...     

Solid state fermentation for production of α-amylase by a thermotolerant Bacillus subtilis from hot-spring water

The production of extracellular α-amylase by thermotolerant Bacillus subtilis was studied in solid state fermentation (SSF). The effect of wheat bran (WB) and rice husk (RH) was examined. The appropriate incubation period, moisture level, particle size and inoculum concentration was determined. Maximum yields of 159,520 and 21,760 U g⁻¹ were achieved by employing WB and RH as substrates in 0.1 M phosphate buffer at pH 7 with 30% initial moisture content at 24 and 48 h. Particle size and inoculum concentration were found to be 1000 μm, 20% and 500 μm, 15% for WB and RH, respectively. Enzyme yield was 7.3-fold higher with WB medium compared with RH.     

Antinoceptive effect of triterpenoid α,β-amyrin in rats on orofacial pain induced by formalin and capsaicin

The effects of α,β-amyrin, a pentacyclic triterpene isolated from Protium heptaphylum was investigated on rat model of orofacial pain induced by formalin or capsaicin. Rats were pretreated with α,β-amyrin (10, 30, and 100 mg/kg, i.p.), morphine (5 mg/kg, s.c.) or vehicle (3% Tween 80), before formalin (20 μl, 1.5%) or capsaicin (20 μl, 1.5 μg) injection into the right vibrissa. In vehicle-treated controls, formalin induced a biphasic nociceptive face-rubbing behavioral response with an early first phase (0–5 min) and a late second phase (10–20 min) appearance, whereas capsaicin produced an immediate face-rubbing (grooming) behavior that was maximal at 10–20 min. Treatment with α,β-amyrin or morphine significantly inhibited the face-rubbing response in both test models. While morphine produced significant antinociception in both phases of formalin test, α,β-amyrin inhibited only the second phase response, more prominently at 30 mg/kg, in a naloxone-sensitive manner. In contrast, α,β-amyrin produced much greater antinociceptive effect at 100 mg/kg in the capsaicin test, which was also naloxone-sensitive. These results provide first time evidence to show that α,β-amyrin attenuates orofacial pain atleast, in part, through a peripheral opioid mechanism but warrants further detailed study for its utility in painful orofacial pathologies.     

Affinity chromatography of α-amylase from Bacillus licheniformis

An affinity chromatographic method with a novel eluant from Bacillus licheniformis is described. α-amylase was bound to starch, starch-celite, starch-Sepharose columns and the bound α-amylase was rapidly eluted with 2% (w/v) white dextrin. The binding capacity of α-amylase to starch column is 380 μmol/g of starch. The purified enzyme showed a single polypeptide on SDS-polyacrylamide gel electrophoresis with a molecular weight of 58 kD. The specificity of purified enzyme was confirmed by immunodiffusion, immunoelectrophoresis. Single radial immunodiffusion and western blotting studies analyzed the synthesis of enzyme at different time points.     

紫草素与依布硒啉对金黄色葡萄球菌的协同作用

[目的]探讨紫草素(shikonin,SKN)协同依布硒啉(ebselen,EbSe)抗金黄色葡萄球菌(Staphylococcus aureus,S.aureus)的作用及机制.[方法]分光光度法测定紫草素-依布硒啉抗金黄色葡萄球菌活性;碘化丙啶(propidium iodide,PI)染色检测紫草素-依布硒啉杀菌效...     

Central and peripheral actions of α-MSH in the thermoregulation of rats

The effect of α-MSH on thermoregulation in rats at room temperature was examined. α-MSH (1 μg ICV or 30 μg IP) alone did not alter temperature. However, this peptide was a potent antipyretic when administered centrally or peripherally in rats treated with pyrogen derived from Salmonella typhi.     

Biotransformation of (±)-α-ionone and β-ionone by cultured cells of Caragana chamlagu

Suspension cultures of Caragana chamlagu (Leguminosae) convert (±)-α-ionone (1) into (±)-3-oxo-α-ionone (3) as the major product and β-ionone (2) into 5,6-epoxy-β-ionone (6) as the sole product. It is interesting to note that the cultured cells of C. chamlagu convert regioselectively the cycloolefinic part of 1 into the corresponding unsaturated carbonyl compound, allylic alcohol and epoxide as the oxidation products, whereas the suspension cultures of Nicotiana tabacum (Solanaceae) convert the unsaturated carbonyl of 1 into the corresponding saturated ketones and alcohols as reduction products.     

Inhibition of chlorophyll biosynthesis by α′,α′-dipyridyl during greening of groundnut leaves

The site of inhibition of chlorophyll biosynthesis by α′,α′-dipyridyl was found to be at the level of conversion of chlorophyllide (672 nm) to chlorophyll (678 nm) during greening of groundnut leaves. This inhibition was partially reversed by certain divalent cations.     

金黄色葡萄球菌病防治研究进展

金黄色葡萄球菌(Staphylococcus aureus, SA)被认为是最常见的食源性致病菌之一,引起人畜的感染性疾病,导致皮肤、软组织和血液感染,引发脓毒症和中毒性休克综合征。随着抗生素的滥用,金黄色葡萄球菌的耐药性逐渐增强,导致耐甲氧西林金黄色葡萄球菌(methicillin resistant Staphylococcus aureus, MRSA)的出现,并且在全球范围内散播,严重危害公共卫生安全。目前亟需有效控制SA感染的新疗法,因此本文对金黄色葡萄球菌防治技术的研究进展进行综述,并对其防治前景进行了分析,以期对金黄色葡萄球菌尤其是MRSA的控制提供理论指导。     

The influence of N-glycosylation on biochemical properties of Amy1, an α-amylase from the yeast Cryptococcus flavus

The yeast Cryptococcus flavus secretes a glycosylated α-amylase (Amy1) when grown in a starch-containing medium. The effects of N-glycosylation on secretion, enzyme activity, and stability of this glycoprotein were studied. Addition of tunicamycin (TM) to the medium at a concentration higher than 0.5 μg mL⁻¹ affected C. flavus growth. Amy1 activity increased by 55% in the intracellular fraction after C. flavus growth in the presence of 0.5 μg mL⁻¹ TM. SDS–PAGE and gel activity detection showed that native enzyme and deglycosylated enzyme had apparent molecular mass of 68 and 64.5 kDa, respectively. The N-glycosylation process did not affect either optimum pH or optimum temperature. The K_M values of native and non-glycosylated α-amylases were 0.052 and 0.098 mg mL⁻¹, and V_max values were 0.038 and 0.047 mg min⁻¹, respectively. However, the non-glycosylated form was more sensitive to inactivation by both the proteolytic enzyme trypsin and high temperature. Furthermore, the activity of the non-glycosylated enzyme was affected by Hg²⁺ and Cu²⁺ suggesting that N-glycosylation is involved in the folding of Amy1.     

15β-hydroxysteroids (Part IV). Steroids of the human perinatal period: The synthesis of 3α,15β,17α-trihydroxy-5α-pregnan-20-one and its A/B-ring configurational isomers

In recent years several 15β-hydroxysteroids have emerged pathognomonic of adrenal disorders in human neonates of which 3α,15β,17α-trihydroxy-5β-pregnan-20-one (2) was the first to be identified in the urine of newborn infants affected with congenital adrenal hyperplasia. In this investigation we report the synthesis of the three remaining 3ξ,5ξ-isomers, namely 3α,15β,17α-trihydroxy-5α-pregnan-20-one (3), 3β,15β,17α-trihydroxy-5α-pregnan-20-one (7) and 3β,15β,17α-trihydroxy-5β-pregnan-20-one (8) for their definitive identification in pathological conditions in human neonates. 3β,15β-Diacetoxy-17α-hydroxy-5-pregnen-20-one (11), a product of chemical synthesis was converted to the isomeric 3 and 7, while conversion of 15β,17α-dihydroxy-4-pregnen-3,20-dione (4), a product of microbiological transformation, resulted in the preparation of 8. In brief, selective acetate hydrolysis of 11 gave 15β-acetoxy-3β,17α-dihydroxy-5-pregnen-20-one (12) which on catalytic hydrogenation gave 15β-acetoxy-3β,17α-dihydroxy-5α-pregnan-20-one (13) a common intermediate for the synthesis of the 3β(and α),5α-isomers. Hydrolysis of the 15β-acetate gave 7, whereas oxidation with pyridinium chlorochromate gave 15β-acetoxy-17α-hydroxy-5α-pregnan-3,20-dione (14) which on reduction with -Selectride and hydrolysis of the 15β-acetate gave 3. Finally, hydrogenation of 4 gave 15β,17α-dihydroxy-5β-pregnan-3,20-dione (10) which on reduction with -Selectride gave 8.     

Characterization of β-endorphin- and α-MSH-related peptides in rat heart

POMC-derived peptides and mRNA have been identified in heart tissue, although POMC processing has not been fully characterized. In the present study, we found that β-lipotropin and ACTH were localized in rat heart, although they were almost entirely converted to β-endorphin- and α-MSH-related peptides. Ion exchange HPLC analysis revealed that β-endorphin(1–31) was further processed to α-N-acetyl-β-endorphin(1–31), which comprised 35.9 ± 0.1% of total immunoreactivity, and smaller amounts of β-endorphin(1–27), β-endorphin(1–26), and their α-N-acetylated derivatives. The predominant α-MSH immunoreactive peptides coeluted with α-MSH and N,O-diacetyl-α-MSH by reverse-phase HPLC, although small amounts of ACTH(1–13)-NH₂ were also present. Thus, multiple forms of β-endorphin and α-MSH are localized in rat heart. β-Endorphin(1–31) is a minor constituent, however, indicating that nonopioid β-endorphin peptides predominate.     

金黄色葡萄球菌tst-1基因敲除菌株的构建

抽提金黄色葡萄球菌834菌株的基因组DNA,PCR克隆扩增tst-1及tst-1的上、下游基因,通过将tst-1上、下游基因分别重组到载体质粒pAULA中,形成同源重组质粒pAULA Δtst-1,将pAULA-Δtst-1电转入细菌内,进行同源重组,以PCR、Western blot鉴定tst-1基因敲除菌株无tst-1基因片段,且无TSST-1蛋白表达,表明已成功构建金黄色葡萄球菌tst-1基因的敲除菌株。