有机介质中脂肪酶催化肌醇烟酸酯的合成

探讨了有机相中固定化脂肪酶催化肌醇和烟酸合成肌醇烟酸酯的可能性；系统地研究了有机溶剂特性、反应初始水活度、温度、pH值、加酶量、底物浓度及摩尔比等因素对酯化反应的影响。     

脂肪酶非水相转化失水山梨醇油酸酯

失水山梨醇单油酸酯（Ｓｐａｎ８０）可在非水相中用脂肪酶酶促转化合成．比较不同来源的脂肪酶和不同的非水相系统后发现：来源于假丝酵母的脂肪酶Ｎｏｖｏｚｙｍ４３５在无溶剂系统中有较高合成Ｓｐａｎ８０的能力．同时考察了水分对酯化反应的影响，并对酯化反应的条件如底物摩尔比、用酶量等进行了研究．与化学合成的产品比较，各项指标基本接近，但酶法合成的产品单酯含量较高，达８０％以上     

有机相酶促合成N-月桂酰-β-氨基丙腈

以月桂酸甲酯和β－氨基丙腈为原料，在有机相中经脂肪酶催化合成N－月桂酰-β-氨基丙腈（LAP）。在比较所用脂肪酶的催化活性的基础上，详细探讨了固定化脂肪酶Candida antarctica（南极洲假丝酵母）CAL－2催化该反应的影响因素。经筛选，适宜的反应条件为：70℃，n（β－氨基丙腈）：n（月桂酸甲酯）＝1：1，底物质量浓度为35g/L，加酶量为200U/g(以氨基丙腈的质量计），反应体系初始加水量为3％腈重。在此实验条件下反应24h后，β－氨基丙腈的最高转化率为96.8%。     

非水相酶促酯交换法合成抗坏血酸脂肪酸酯

研究了一种L-抗坏血酸脂肪酸酯（AFAE）合成新途径。实验结果表明，氢化棕榈油、大豆油和海狗油等食用油脂作为脂肪酸基团供体，在固定化脂肪酶的催化下，在有机溶荆中与L-抗坏血酸直接进行酯交换反应形成L-抗坏血酸脂肪酸酯。对反应介质体系的筛选发现，在叔戊醇中用Novozym435催化此反应所得的产物浓度最高。对影响合成抗坏血酸脂肪酸酯的因素进行了探讨，确定了最适反应条件：初始油脂底物浓度为200-600mmol／L，温度为55℃，反应时间为9h。此条件下产物质量浓度可达43．51g／L。     

固定化脂肪酶Lipozyme TL IM催化菜籽油醇解反应合成生物柴油的工艺条件

脂肪酶Lipozyme TL IM在有机相体系中能有效催化菜籽油醇解反应生成生物柴油。作者研究了酶量、底物比率、反应时间、反应温度、转速、水分质量分数、有机溶剂对产物得率的影响。最佳工艺条件为：在正己烷的反应介质中,酶量50%,醇油摩尔比3∶1,反应温度50℃,转速150r/min,添加质量分数5%的水分,反应12 h后产物脂肪酸甲酯得率可达92.3%。     

无溶剂体系酶促合成甘油中碳酸偏酯

根据中碳酸转化率的大小 ,对 5种不同来源的固定化脂肪酶筛选 ,其中固定化脂肪酶SSW的酶催化转化率最高 ,辛酸和癸酸转化率分别为 91.5 %和 93.5 % ;对脂肪酶SSW酶促合成甘油辛酸偏酯的反应条件优化 ,适宜反应条件为 :敞口反应器中反应温度 5 5℃ ,每克酸加酶量 10 0U/g ,辛酸 /甘油投料摩尔比 1∶1.1,甘油初始加水量 4 % (质量分数 ) .实验范围内中碳酸品种对脂肪酶SSW不显示基质特异性 .     

微乳凝胶固定化酶催化长链脂肪酸酯的合成

研究了以AOT(二 (2 乙基已基 )琥珀酸酯磺酸钠 ) /正庚烷 /水 /明胶组成的反相微乳凝胶固定化脂肪酶 ,在有机溶剂中催化棕榈酸和十六醇的酯化反应 .反应在 2 5℃下进行 ,固定化脂肪酶重复使用 16次后 ,平衡转化率仍可达 90 % .反应既可用釜式反应器又适用柱式反应器 .产物通过冷却的反应液后沉淀析出 ,回收率可达 6 5% ,产品纯度高 .     

有机相中微生物脂肪酶合成短链脂肪酸酯的研究

用微生物脂肪酶在溶剂相中合成短链脂肪酸酯的研究表明：脂肪酶在正庚烷中催化合成短链脂肪酯转化率比水相中有明显的优势。在１０个不同来源的商品脂肪酶中，来自Ｍｕｃｏｒｍｉｅｈｅｉ，Ｃａｎｄｉｄａｒｕｇｏｓａ和Ｐｏｒｃｉｎｅｐａｎｃｒｅａｓ的脂肪酶合成酯转化率较高，其中Ｍｕｃｏｒｍｉｅｈｅｉ脂肪酶对己酸乙酯４８ｈ合成转化率达９４％，己酸乙酯产量３３．８４ｇ／Ｌ．同时就底物酸和醇的碳链长度对酯转化率的影响进行了研究。     

利用胆固醇氧化酶转化胆固醇制备胆甾-4-烯-3-酮

用酶法催化氧化胆固醇制备胆甾-4-烯-3-酮相对于化学法合成具有反应简单、成本较低等优点.作者探讨了在正辛烷作为有机相的两相反应体系中,以胆固醇氧化酶催化氧化胆固醇制备胆甾-4-烯-3-酮的方法.在胆固醇质量浓度为40g/L,反应时间40min、反应温度40℃、磷酸缓冲液-正辛烷体积比32、通氧40L/h及搅拌转速300r/min下,胆固醇转化率达到92%,经薄板层析及紫外扫描,发现转化产物为单一的胆甾-4-烯-3-酮.     

温度对单甘酯的合成起关键作用

在无溶剂体系中以脂肪酶催化棕榈油的甘油解反应合成单甘酯时，发现温度对单甘酯的产率起关键作用。反应存在一临界温度，低于临界温度时单甘酯的平衡浓度高于６０％，而高于临界温度时单甘酯含量只有３０％左右。临界温度与油脂的种类有关；通过测定各种油脂的临界温度并与其熔点相比较，结合反应现象学及反应进程中反应混合物各组分含量的变化，提出了临界温度的存在机理。     

Activation of serum complement inhibits collagen synthesis in fetal rat bone in organ culture

Summary Activation of rabbit serum complement caused a marked reduction in collagen synthesis but a much smaller change in noncollagen protein synthesis in fetal rat calvaria maintained in organ culture. In the periosteum of the fetal rat calvarium, both collagen and noncollagen protein synthesis were reduced, whereas in the central bone, presumably enriched in osteoblasts, only collagen synthesis was inhibited. This large decrease in bone collagen synthesis could not be attributed to enhanced degradation of newly synthesized collagen or its release into the culture medium. Activation of complement also stimulated the production of PGE in fetal rat calvaria. Antagonists of prostaglandin cyclooxygenase decreased prostaglandin synthesis but did not restore collagen synthesis in complement-treated bones, suggesting that complement decreases osteoblast collagen synthesis by a mechanism largely independent of prostaglandin production.     

Activation of serum complement inhibits collagen synthesis in fetal rat bone in organ culture

Activation of rabbit serum complement caused a marked reduction in collagen synthesis but a much smaller change in noncollagen protein synthesis in fetal rat calvaria maintained in organ culture. In the periosteum of the fetal rat calvarium, both collagen and noncollagen protein synthesis were reduced, whereas in the central bone, presumably enriched in osteoblasts, only collagen synthesis was inhibited. This large decrease in bone collagen synthesis could not be attributed to enhanced degradation of newly synthesized collagen or its release into the culture medium. Activation of complement also stimulated the production of PGE in fetal rat calvaria. Antagonists of prostaglandin cyclooxygenase decreased prostaglandin synthesis but did not restore collagen synthesis in complement-treated bones, suggesting that complement decreases osteoblast collagen synthesis by a mechanism largely independent of prostaglandin production.     

Hepatic acute phase protein synthesis is indirectly regulated by tumor necrosis factor.

It has been proposed that tumor necrosis factor (TNF) is a direct regulator of postinjury hepatic protein synthesis. To test this hypothesis we investigated the total protein and specific acute phase protein synthesis response of murine hepatocytes to stimulation with mu-rTNF-alpha in vivo and in vitro. Total hepatocyte secretory protein synthesis was assessed by incorporation of [35-S] methionine into TCA-precipitated protein; and acute phase protein synthesis was assessed by induction of a 23-kD acute phase protein marker and by suppression of albumin synthesis determined by SDS-PAGE and autoradiography. We found that rTNF in vivo (8,000 units, IP injection) was associated with reduced total hepatocyte secretory protein synthesis (29 +/- 10%), increased synthesis of the 23-kD acute phase reactant (4.1 +/- 1.6-fold), and decreased albumin synthesis (0.68 +/- 0.2-fold) compared to saline-injected control animals. The in vitro stimulation of cultured murine hepatocytes directly with rTNF failed to demonstrate changes in total secretory protein synthesis or 23-kD protein; however, it did result in significant suppression of albumin synthesis (0.82 +/- 0.1-fold). In additional experiments, hepatocytes:nonparenchymal cell co-cultures stimulated with lipopolysaccharide (LPS) demonstrated protein synthesis changes similar to the in vivo TNF response including increased 23-kD protein and decreased albumin synthesis. These co-cultures demonstrated TNF production; however, addition of TNF antiserum during LPS stimulation had no effect on either 23-kD protein or albumin synthesis, despite the complete neutralization of TNF activity in the co-culture supernatants.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cerivastatin inhibits fetal calf serunvinduced DNA synthesis in cultured rat mesangial cells

SUMMARY: Inhibition of mevalonate synthesis by several statins has been shown to suppress DNA synthesis in glomerular mesangial cells. In the present study, we investigated the effect of a new statin, cerivastatin, on fetal calf serum (FCS)-induced DNA synthesis of cultured rat mesangial cells. Cultured rat mesangial cells were stimulated by 10% FCS in the presence or absence of cerivastatin and mevalonate. 5-bromo-2-deoxyuridine (BrdU) incorporation was used to assess DNA synthesis. the present study showed that 10% FCS caused marked stimulation of DNA synthesis in the mesangial cells. Cerivastatin inhibited FCS-stimulated BrdU incorporation in a dose-dependent manner. IC₅₀ was approximately 1 umol/L. Exogenous mevalonate, farnesyl pyrophosphate and geranylgeranyl pyrophosphate significantly prevented the inhibitory effect of cerivastatin on DNA replication. It appears that cerivastatin, by inhibiting the synthesis of mevalonate, may suppress DNA synthesis in the mesangial cells.     

抑癌基因MTS1对人瘢痕疙瘩成纤维细胞的生物学影响

目的：探讨多肿瘤抑制基因1（MTS1）对人瘢痕疙瘩成纤维细胞在生长增殖、DNA合成代谢及胶原合成量等方面的影响。方法：将外源性MTS1基因导入人瘢痕疙瘩成纤维细胞后，通过MTT法测定细胞的生长曲线，通过^3H-胸腺嘧啶核苷（^3H-TdR）及^3H-脯氨酸掺入的方法测定细胞的DNA合成量及胶原合成量，来分别比较转染前后细胞的生物学变化。结果：在转染MTS1后瘢痕疙瘩成纤维细胞的生长增殖受到明显的抑制，同时其DNA合成量及胶原合成量也明显降低，而转染空载体组及正常组细胞均未出现上述变化。结论：外源性多肿瘤抑制基因1(MTS1）能明显抑制人瘢痕疙瘩成纤维细胞在生长增殖、DNA合成及胶原合成等方面生物学功能，为瘢痕疙瘩的临床治疗提供了新的思路。     

Inhibition of bone collagen synthesis by the tumor promoter phorbol 12-myristate 13-acetate

We characterized the effect of the tumor promoter phorbol 12-myristate 13-acetate (PMA) on osteoblast function and DNA synthesis in 21-day-old fetal rat calvaria maintained in organ culture. Protein synthesis was determined by measuring the incorporation of [3H]proline into collagenase-digestible (CDP) and noncollagen protein (NCP), respectively. Alkaline phosphatase activity was assessed as the release of p-nitrophenol from p-nitrophenol phosphate. DNA synthesis was determined by the incorporation of [3H]thymidine into acid-insoluble bone and total DNA content. PMA at 3-100 ng/ml (4-133 nM) caused a dose-related inhibition of collagen synthesis that was observed 6 hours after adding PMA to calvaria. PMA inhibited collagen synthesis in the osteoblast-rich central bone of calvaria but did not alter collagen synthesis in the periosteum. There was little effect of PMA on noncollagen protein synthesis in the central bone or periosteum. Phorbol esters that do not promote tumor formation in vivo did not alter collagen synthesis in calvaria. PMA stimulated prostaglandin E2 (PGE2) production in calvaria, but indomethacin did not alter the inhibitory effect of PMA on bone collagen synthesis. PMA decreased alkaline phosphatase activity measured after 48 hr of culture and increased the incorporation of [3H]thymidine into bone and DNA content after 96 hr of culture. These data indicate that PMA inhibits collagen synthesis and alkaline phosphatase activity, while stimulating DNA synthesis, suggesting that activation of protein kinase C might regulate osteoblast function and bone cell replication.     

In vitro DNA synthesis in freshly separated human breast cancer cells assessed by tritiated thymidine incorporation assay: relationship to the long-term outcome of patients

BACKGROUND: Tumour growth rate has a significant effect on the clinical course of various malignancies. The present study was designed to assess whether in vitro DNA synthesis in freshly separated breast cancer cells is a useful marker in evaluating growth rates and in predicting the clinical outcome of patients. METHODS: From 1982 to 1992, DNA synthesis was assessed by [3H]thymidine incorporation in 97 samples of primary lesions from 94 patients with breast cancer. The patients were followed for 5-15 years and their outcome was surveyed in January 1998. RESULTS: The level of DNA synthesis did not correlate with the patients' age, clinical stage or expression of oestrogen receptor. However, it correlated significantly with the histological grade. In 89 patients, whose outcome was reported, the survival rate in the group with a high rate of DNA synthesis (log10c.p.m. 3.0 or more) was significantly lower than that in the low-level group; the 5- and 10-year survival rates were 84 and 74 per cent for the low synthesis group (n = 46), and 60 and 46 per cent for the high synthesis group (n = 43) respectively. This was also noted in patients with stage 1 or 2 cancers, for whom the 5- and 10-year survival rates were 100 and 90 per cent for the low synthesis group (n = 25), and 75 and 70 per cent for the high synthesis group (n= 35). Multivariate analysis supported this significant correlation for DNA synthesis in the prognosis of patients after mastectomy. Furthermore, the level of DNA synthesis was significantly higher in 18 patients who died from a recurrence within 3 years after operation than in 56 survivors, and the level of DNA synthesis also correlated significantly with the survival period in the 33 patients who died. CONCLUSION: The level of DNA synthesis in breast cancer was variable, and was independent of the clinical stage or oestrogen receptor status. However, a high level of DNA synthesis was a positive indicator of a high risk of recurrence after operation, especially for stage 1 or 2 breast cancer. In vitro DNA synthesis may account for some of the clinical characteristics of breast cancers.     

Activities of a cancellous bone-cell-stimulating substance

A bone-cell-stimulating substance (BCSS) that initiates appositional bone formation in intact rats was examined for its effects on DNA and collagen synthesis in tibial and calvarial organ cultures of 17-day-old embryonic chicks. BCSS stimulated collagen synthesis in both types of bone. BCSS stimulated DNA synthesis in tibiae but inhibited synthesis in calvaria from the same chicks. Insulinlike growth factor, transforming growth factor-beta, platelet-derived growth factor, and fetal bovine serum also affected DNA synthesis differently in calvaria and tibiae. BCSS was able to modify some of the effects of these growth factors on DNA.     

The effect of cholesterol feeding and alterations in bile acid homeostasis on de novo sterologenesis in diabetic rats

Previous studies have demonstrated that de novo cholesterol synthesis is increased two- to threefold in the intestines of diabetic animals. This increase is due to a stimulation of cholesterogenesis in both the small and large intestine but, quantitatively, the small intestine is primarily responsible for the observed increase. The present study examined the effect of cholesterol feeding and alterations of bile acid homeostasis on de novo sterol synthesis in intact normal and diabetic animals. Cholesterol feeding in the control animals did not affect sterol synthesis in the small intestine, but in diabetic animals cholesterol feeding markedly inhibited small intestinal sterologenesis. The threefold stimulation of small intestinal sterol synthesis observed in diabetic animals is completely obliterated by cholesterol ingestion. Moreover, this inhibition of sterol synthesis by cholesterol feeding in the small intestine of diabetic animals occurred very rapidly (within 36 h). In the large intestine, cholesterol feeding did not influence sterol synthesis in either the diabetic or control animals. In the liver, cholesterol feeding markedly inhibited sterol synthesis to similar degrees in the diabetics and controls. Colestipol feeding and biliary drainage, procedures that reduce bile acid pool size, stimulated sterol synthesis in the liver and small intestine of both diabetic and control animals. However, reductions in bile acid pool size increased sterologenesis in the large intestine in control animals but had no effect in the diabetics. Bile acid ingestion did not alter either small or large intestinal sterologenesis in the diabetic or control animals. In conclusion, the present study demonstrates the sterol synthesis is enhanced in the small and large intestine of diabetic animals and, moreover, both the cholesterol- and bile acid-mediated regulation of cholesterol synthesis in the intestines of the diabetic animals is altered from normal.     

Fibrin monomer and fibrinopeptide B act additively to increase DNA synthesis in smooth muscle cells cultured from human saphenous vein

PURPOSE: We investigated the hypothesis that fibrinogen increased DNA synthesis (and cell proliferation) of smooth muscle cells (SMCs) cultured from human saphenous vein and that the increased DNA synthesis was attenuated when cells were cultured on polymeric collagen. METHODS: SMCs were cultured from human saphenous vein on plastic, fibronectin, monomeric, and polymeric collagen. Fibrinogen products were prepared by proteolytic digestion. DNA synthesis was measured by bromodeoxyuridine incorporation into DNA, cell proliferation by cell counting, cyclic adenosine monophosphate by enzyme-linked immunosorbent assay, and fibrinopeptide B labeled with iodine 125 used for binding studies. RESULTS: Fibrin monomer (0.003-0.1 micromol/L) stimulated a concentration-dependent increase in DNA synthesis of up to 10-fold, which could be inhibited by the peptide Bbeta15-42. The stimulation of DNA synthesis was highest for cells cultured on plastic and lowest for cells cultured on type I collagen polymer. Much higher concentrations of fibrinogen (0.3-1 micromol/L) were required to effect similar increases in DNA synthesis. Fibrinogen had a particular effect to augment DNA synthesis, up to 14-fold, when cells were cultured on monomeric type I collagen. This augmented DNA synthesis was inhibited by a neutralizing antibody to urokinase-type plasminogen activator. Incubation of cells cultured on collagen monomer with fibrinogen resulted in production of fibrinopeptide B. Fibrinopeptide B (5 micromol/L) increased DNA synthesis by fourfold and had additive effects with fibrin monomer to increase DNA synthesis. Iodinated tyrosine fibrinopeptide B bound to SMCs (dissociation constant 0.6 micromol/L). CONCLUSION: Cultured human saphenous vein SMCs appear to have high-affinity receptors for fibrin monomer and fibrinopeptide B, the engagement of which stimulates DNA synthesis. These mechanisms may be pertinent to the association between fibrinogen and vein graft stenosis in vivo.