Total synthesis of bis-(2,3-dibromo-4,5-dihydroxyphenyl)-methane as potent PTP1B inhibitor

Protein tyrosine phosphatase 1B (PTP1B) plays an important role as a negative regulator and has been proved to be an effective target for the treatment of type 2 diabetes mellitus. Bis-(2,3-dibromo-4,5-dihydroxyphenyl)-methane 7 was first reported as a natural bromophenol with significant inhibition against PTP1B which was isolated from red algae Rhodomela confervoides. Intrigued by its astonishing activity (IC50 = 2.4 mu mol/L), compound 7 was synthesized with the overall yield of 24% and evaluated for its PTP1B inhibitory activity compared with natural compound. (C) 2008 Li Jun Han. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.

1-（2-萘基）-3-甲基-5-吡唑啉酮衍生试剂的制备及其在高效液相色谱-质谱法测定糖类化合物中的应用

以1-（2-萘基）-3-甲基-5-吡唑啉酮（NMP）作为柱前衍生试剂,建立了简单、灵敏的糖类组分的反相高效液相色谱测定方法。NMP与糖在氨为催化剂的条件下,于70℃下反应可获得稳定的衍生产物。在HypersilODS2反相色谱柱上,实现了8种单糖的基线分离。衍生物线性相关系数均大于0.9985,检出限为0.58-1.1pmol。利用柱后在线串联质谱的电喷雾电离正离子模式监测,获得了各组分的质谱定性及裂解规律,特别是m/z473的特征碎片离子可作为单糖NMP衍生物的判定依据。与1-苯基-3-甲基-5-吡唑啉酮（PMP）相比,NMP对糖的衍生化具有灵敏、简单、质谱裂解规律性强、重现性好等优点。该方法用于测定油菜花粉多糖中的单糖组成,结果令人满意。

植物群落生物量和有机碳对高寒草甸土地利用变化的响应

在中国科学院海北高寒草甸生态系统定位站地区,选择高寒矮嵩草草甸及其开垦后形成的农田和一年生人工草地作为研究对象,研究了高寒草甸不同土地利用方式下生物量和植物-土壤系统固定的有机碳量的变化。结果表明：3种土地利用方式相比较,地上生物量由高到低依次为人工草地〉农田〉高寒草甸（P〈0.01）,分别为11.83、9.78和4.36 t/hm~2;3种土地利用方式下地下生物量剖面分布均呈倒金字塔形,0～40 cm地下生物量为高寒草甸〉人工草地〉农田（P〈0.01）,分别为15.74、5.61和1.24 t/hm~2。随着高寒草甸土地利用方式改变,植物群落碳素固定量也随之减小,其序列由高到低依次为：高寒草甸〉人工草地〉农田（P〈0.05）,其值分别为7.63、6.81和4.51 t/hm~2。

石漠化过程中土壤——植被系统营养元素协变关系

贵州位于世界上连片分布面积最大的中国西南岩溶区的中心，近年来石漠化问题突出，且有不断恶化的趋势，给当地经济发展和人类生存带来了极大障碍。对石漠化的成因机理进行研究，是石漠化治理的前提和理论依据。但目前对石漠化的研究基本以定性分析为主，定量化不足。没有统一的石漠化等级划分标准， 对于石漠化过程土壤、植物的退化本质也不是很清楚，这给石漠化治理带来了极大的障碍。 本文通过在小流域尺度上，以贵州花江峡谷查耳岩小流域为研究区域，对喀斯特石漠化过程土壤、植被退化进行定量分析,以确定土壤－植被系统营养元素协变关系，分析了石漠化演替过程中土壤、植被的变化，土壤、植被间关联退化的关系。还检验了景观上石漠化划分方法是否能体现出退化的本质。取得了以下几点认识： (1) 在研究区土壤pH值在6.9-7.8的中性偏碱性范围,土壤总钙、交换态钙含量高，氮含量丰富，有机质、养分全量较高，有效态含量除了速效K、P在部分样地含量较低外，其它营养元素的有效态养分含量都较高。 (2) 在喀斯特石漠化地区植物灰分含量高，富钙、氮，相对缺铁、锌、钾、磷，植物灰分含量与植物钙含量成显著正相关，即随着植物钙含量的增加植物灰分含量增加。 (3) 通过对四套石漠化序列比较得出，在弃耕地两套石漠化序列中，土壤为棕色石灰岩土，土层较厚，土壤覆盖度、总量相对较高，但是有机质、有效态养分含量相对低，随着演替土壤质量有下降趋势，但是没有明显规律。樵采的两套石漠化序列，土壤为发育较年轻的黑色石灰土，土层很薄，土壤覆盖度低，总量少，零星分布在负地形中，但是土壤有机质、全量养分、速效养分含量都较丰富，随着石漠化演替没有呈现相应的规律变化。 (4) 按照目前景观上石漠化的划分标准，在土壤退化上，没有表现出随石漠化演替土壤质量发生规律退化。这主要由于喀斯特山地的特殊性，在石灰土地区土壤厚度差异很大，分布零星，在土壤很薄的地方只要发生轻微的水土流失可能会出现强度石漠化的景观，而在土壤相对厚的地方要经过较强的水土流失才会出现石漠化景观。尽管开垦和樵采两种不同的人为干扰方式下产生的石漠化景观可能很相似，但是在土壤退化程度上不一致，而景观划分标准不能区分这两种情况，导致了土壤并没有表现出随石漠化演替出现规律退化。要将土壤退化这一本质内容涉及到石漠化等级划分中，应将土壤质量评价方面的因子及其量化标准补充到石漠化划分标准中。 (5) 研究区植被在结构上与石漠化的演替形成很好对应关系，随着石漠化的演替，植被从乔－草灌－灌草－稀疏灌草演化，但是植物营养元素、灰分没有随着石漠化的演替而发生规律变化，表明在景观上的划分标准体现了植被宏观的变化，但是对于植物营养元素、灰分含量的微观变化没有呈现出相应的变化规律。 (6) 土壤元素有效态含量与土壤pH值、机械组成的相关性弱或者不相关，与养分全量、有机质的相关性较强。 (7)石漠化过程中，土壤——植物系统表现出复杂的关系。土壤各项物理性质及元素的全量、有效态含量与植物灰分、元素含量的相关性很弱或者基本不相关，随着石漠化的演替土壤——植物系统营养元素也没有表现出规律变化。

Ligation of cell surface GRP78 with antibody directed against the COOH-terminal domain of GRP78 suppresses Ras/MAPK and PI 3-kinase/AKT signaling while promoting caspase activation in human prostate cancer cells.

We have previously shown that treatment of prostate cancer and melanoma cells expressing GRP78 on their cell surface with antibody directed against the COOH-terminal domain of GRP78 upregulates and activates p53 causing decreased cell proliferation and upregulated apoptosis. In this report, we demonstrate that treatment of 1-LN prostate cancer cells with this antibody decreases cell surface expression of GRP78, Akt(Thr308) and Akt(Ser473) kinase activities and reduces phosphorylation of FOXO, and GSK3beta. This treatment also suppresses activation of ERK1/2, p38 MAPK and MKK3/6; however, it upregulates MKK4 activity. JNK, as determined by its phosphorylation state, is subsequently activated, triggering apoptosis. Incubation of cells with antibody reduced levels of anti-apoptotic Bcl-2, while elevating pro-apoptotic BAD, BAX and BAK expression as well as cleaved caspases-3, -7, -8 and -9. Silencing GRP78 or p53 gene expression by RNAi prior to antibody treatment abrogated these effects. We conclude that antibody directed against the COOH-terminal domain of GRP78 may prove useful as a pan suppressor of proliferative/survival signaling in cancer cells expressing GRP78 on their cell surface.

Chemoenzymatic synthesis of enantiopure dihydroxy-1,2,3,4-tetrahydronaphthalenes from naphthalene and dihydronaphthalene precursors

cis-Dihydrodiol, cis-tetrahydrodiol and arene hydrate bacterial metabolites, of naphthalene and 1,2-dihydronaphthalene, have been used as synthetic precursors; chemoenzymatic and enzyme-catalysed syntheses have been used to obtain all possible enantiopure samples of dihydroxy-1,2,3,4-tetrahydronaphthalene stereoisomers.

Kinetic analysis of the cleavage of human protease-activated receptor-1/2/3 and 4 using quenched-fluorescent peptide substrates

Protease-activated receptors [PARs] are a family of G-protein-coupled seven-transmembrane domain receptors that are activated by proteolytic cleavage of their amino-terminal exodomain. To characterize the cleavage rate of human PAR-1 / 2 / 3 and 4 by trypsin and thrombin, four synthetic quenched-fluorescent peptide substrates have been synthesized. Each substrate consisted of a ten-residue peptide spanning the receptor activation cleavage site and using progress-curve kinetics, k(cat)/K-m values were determined.

A Dual Role for Lecithin:Cholesterol Acyltransferase (EC 2.3.1.43) in Lipoprotein Oxidation

Lecithin:cholesterol acyltransferase (LCAT) is a key enzyme involved in lipoprotein metabolism. It mediates the transesterification of free cholesterol to cholesteryl ester in an apoprotein A-I-dependent process. We have isolated purified LCAT from human plasma using anion-exchange chromatography and characterized the extracted LCAT in terms of its molecular weight, molar absorption coefficient, and enzymatic activity. The participation of LCAT in the oxidation of very low density lipoproteins (VLDL) and low-density lipoproteins (LDL) was examined by supplementing lipoproteins with exogenous LCAT over a range of protein concentrations. LCAT-depleted lipoproteins were also prepared and their oxidation kinetics examined. Our results provide evidence for a dual role for LCAT in lipoprotein oxidation, whereby it acts in a dose-responsive manner as a potent pro-oxidant during VLDL oxidation, but as an antioxidant during LDL oxidation. We believe this novel pro-oxidant effect may be attributable to the LCAT-mediated formation of oxidized cholesteryl ester in VLDL, whereas the antioxidant effect is similar to that of chain-breaking antioxidants. Thus, we have demonstrated that the high-density lipoprotein-associated enzyme LCAT may have a significant role to play in lipoprotein modification and hence atherogenesis. (C) 2007 Elsevier Inc. All rights reserved.

Three component coupling reactions of N-acetyl-2-azetine-rapid stereoselective entry to 2,3,4-trisubstituted tetrahydroquinolines

N-Acetyl-2-azetine undergoes Lewis acid catalysed [4 + 2]-cycloaddition with imines derived from aromatic amines and gave a 1:1 mixture of exo-endo diastereoisomeric azetidine cycloadducts which reacted further with aromatic amine, to give 2,3,4-trisubsitituted tetrahydroquinolines in good to excellent yield, predominantly as one diastereoisomer.

Primed, constant infusion with [H-2(3)]serine allows in vivo kinetic measurement of serine turnover, homocysteine remethylation, and transsulfuration processes in human one-carbon metabolism

Background: One-carbon metabolism involves both mitochondrial and cytosolic forms of folate-dependent enzymes in mammalian cells, but few in vivo data exist to characterize the biochemical processes involved.

Objective: We conducted a stable-isotopic investigation to determine the fates of exogenous serine and serine-derived one carbon units in homocysteine remethylation in hepatic and whole-body metabolism.

Design: A healthy man aged 23 y was administered [2,3,3 H-2(3)]serine and [5,5,5-H-2(3)]leucine by intravenous primed, constant infusion. Serial plasma samples were analyzed to determine the isotopic enrichment of free glycine, serine, leucine, methionine, and cystathionine. VLDL apolipoprotein B-100 served as an index of liver free amino acid labeling.

Results: [H-2(1)]Methionine and [H-2(2)]methionine were labeled through homocysteine remethylation. We propose that [H-2(2)]methionine occurs by remethylation with [H-2(2)]methyl groups (as 5-methyltetrahydrofolate) formed only from cytosolic processing of [H-2(3)]serine, whereas [H-2(1)]methionine is formed with labeled one-carbon units from mitochondrial oxidation of C-3 serine to [H-2(1)]formate to yield cytosolic [H-2(1)]methyl groups. The labeling pattern of cystathionine formed from homocysteine and labeled serine suggests that cystathionine is derived mainly from a serine pool different from that used in apolipoprotein B-100 synthesis.

Conclusions: The appearance of both [H-2(1)]- and [H-2(2)]methionine forms indicates that both cytosolic and mitochondrial metabolism of exogenous serine generates carbon units in vivo for methyl group production and homocysteine remethylation. This study also showed the utility of serine infusion and indicated functional roles of cytosolic and mitochondrial compartments in one-carbon metabolism.

Toluene dioxygenase-catalyzed cis-dihydroxylation of benzo[b]thiophenes and benzo[b]furans: synthesis of benzo[b]thiophene 2,3-oxide

Enzymatic cis-dihydroxylation of benzo[b]thiophene, benzo[b]furan and several methyl substituted derivatives was found to occur in both the carbocyclic and heterocyclic rings. Relative and absolute configurations and enantiopurities of the resulting dihydrodiols were determined. Hydrogenation of the alkene bond in carbocyclic cis-dihydrodiols and ring-opening epimerization/reduction reactions of heterocyclic cis/trans-dihydrodiols were also studied. The relatively stable heterocyclic dihydrodiols of benzo[b]thiophene and benzo[b]furan showed a strong preference for the trans configuration in aqueous solutions. The 2,3-dihydrodiol metabolite of benzo[b]thiophene was utilized as a precursor in the chemoenzymatic synthesis of the unstable arene oxide, benzo[b]thiophene 2,3-oxide.

Decrease in skin collagen glycation with improved glycemic control in patients with insulin-dependent diabetes mellitus

Glycation, oxidation, and nonenzymatic browning of protein have all been implicated in the development of diabetic complications. The initial product of glycation of protein, fructoselysine (FL), undergoes further reactions, yielding a complex mixture of browning products, including the fluorescent lysine-arginine cross-link, pentosidine. Alternatively, FL may be cleaved oxidatively to form N(epsilon)-(carboxymethyl)lysine (CML), while glycated hydroxylysine, an amino-acid unique to collagen, may yield N(epsilon)-(carboxymethyl)hydroxylysine (CMhL). We have measured FL, pentosidine, fluorescence (excitation = 328 nm, emission = 378 nm), CML, and CMhL in insoluble skin collagen from 14 insulin-dependent diabetic patients before and after a 4-mo period of intensive therapy to improve glycemic control. Mean home blood glucose fell from 8.7 +/- 2.5 (mean +/- 1 SD) to 6.8 +/- 1.4 mM (P less than 0.005), and mean glycated hemoglobin (HbA1) from 11.6 +/- 2.3% to 8.3 +/- 1.1% (P less than 0.001). These changes were accompanied by a significant decrease in glycation of skin collagen, from 13.2 +/- 4.3 to 10.6 +/- 2.3 mmol FL/mol lysine (P less than 0.002). However, levels of browning and oxidation products (pentosidine, CML, and CMhL) and fluorescence were unchanged. These results show that the glycation of long-lived proteins can be decreased by improved glycemic control, but suggest that once cumulative damage to collagen by browning and oxidation reactions has occurred, it may not be readily reversed. Thus, in diabetic patients, institution and maintenance of good glycemic control at any time could potentially limit the extent of subsequent long-term damage to proteins by glycation and oxidation reactions.