高产黑色素微生物资源的研究

对文献报道的产黑色素的八株微生物进行了比较研究,从比较中发现嗜麦芽假单胞菌Ｐ９和链霉菌１５４５产黑色素能力较强,并从土壤中初筛得到１３株产色素菌株。选取产黑色素较高的Ｔ１,Ｔ１１两株菌进行比较研究。研究结果表明,在ｐＨ５．５的发酵培养基中,振荡培养（转速２８０ｒｐｍ,３０℃）的Ｔ１菌株产生的可溶性黑色素最高,其ＯＤ４００可达０．５８１×１０。此外还对Ｔ１菌株的一些特性进行了初步研究。     

一株高产黑色素香灰菌菌株的鉴定、筛选及培养条件的优化

为筛选一株产黑色素能力强的菌株并优化其培养条件。通过ITS测序鉴定11株供试菌株,以菌丝生长速度、平板L值等指标筛选出一株产黑能力强的香灰菌,并对其生长所需碳源、氮源、pH等培养条件进行优化。研究表明,11株供试菌株均为香灰菌（Hypoxylon sp.）,其中Hp.sp0006菌丝生长速度较快、菌球大且均匀、L值最低,并且发酵液黑色素含量最高。该菌株最优培养条件是,以葡萄糖为碳源、牛肉浸膏为氮源、碳氮比20∶1并添加10 mg维生素B₁,黑色素含量可达（1.21±0.17）g/L。香灰菌Hp.sp0006是一株产黑色素较高的菌株,优化后的培养基更有利于黑色素的合成,为香灰菌黑色素的开发利用奠定基础。     

嗜热菌Anoxybacillu sp.产淀粉酶培养基优化及产酶条件研究

目的：对产淀粉酶嗜热菌Anoxybacillu sp.菌株进行培养基优化及产酶条件研究,以便提高菌株的产酶能力,并为下一步菌 株的诱变育种研究提供基础。方法：常规方法液体培养菌株,用平板初筛和DNS法复筛选择产淀粉酶能力较高的菌株;单因素筛 选培养基最适的碳源、氮源、Ca2+浓度和Mg2+浓度,对单因素筛选的最佳碳源、氮源、Ca2+和Mg2+的三个较佳浓度进行四因素三 水平正交试验优化培养基;对培养基不同pH值及不同培养温度进行培养条件研究。结果：产淀粉酶菌株筛选结果显示：六株菌中 淀粉酶酶活力值最大的是菌株Anoxybacillu sp.DL4,差异有统计学意义（P＜0.05）。培养基单因素筛选结果显示：最适碳源为麦芽糖、最适氮源为 硝酸铵、最适Ca2+、Mg2+浓度均为0.02%,差异有统计学意义（P＜0.05）。培养基优化结果显示：C 源0.1 %,N源0.2 %,Mg2+ 0.04%, Ca2+ 0.04 %为最佳的培养基成分组合。产酶条件筛选结果显示：培养基pH 值为6、培养温度为55 ℃时菌株产酶水平最高,差异有 统计学意义（P＜0.05）。结论：培养基的优化及最适的产酶条件能提高嗜热菌Anoxybacillu sp.DL4 产淀粉酶能力,Ca2+、Mg2+离子 对菌株产淀粉酶有促进作用。     

CN—92植酸酶产生菌的诱变选育及产酶条件的研究

以无花果曲霉IFFI2227为出发菌株,通过NTGT和Co60复合诱变处理,获得5株产植酸酶活力比出发菌株高2．1～2．5倍的高产菌株。其中1株CN－92菌产酶性能稳定,研究了该菌株的最适产酶条件：培养温度30℃,培养基起始pH6．0培养时间72h。适宜于该菌株产植酸酶的优良固体培养基组成为：麸皮、豆饼粉、硫酸铵及少量无机磷,培养基含水率51％。葡萄糖、乳糖和MH4Cl、NH4NO3等不同程度地抑制植酸酶的合成,微量元素如Mn,Zn,Cu等对产酶无促进作用。     

嗜热菌Anoxybacillus sp.产淀粉酶培养基优化及产酶条件研究

目的:对产淀粉酶嗜热菌Anoxybacillus sp.菌株进行培养基优化及产酶条件研究,以便提高菌株的产酶能力,并为下一步菌株的诱变育种研究提供基础。方法:常规方法液体培养菌株,用平板初筛和DNS法复筛选择产淀粉酶能力较高的菌株;单因素筛选培养基最适的碳源、氮源、Ca~(2+)浓度和Mg~(2+)浓度,对单因素筛选的最佳碳源、氮源、Ca~(2+)和Mg~(2+)的三个较佳浓度进行四因素三水平正交试验优化培养基;对培养基不同p H值及不同培养温度进行培养条件研究。结果:产淀粉酶菌株筛选结果显示:六株菌中淀粉酶酶活力值最大的是菌株DL4,差异有统计学意义(P0.05)。培养基单因素筛选结果显示:最适碳源为麦芽糖、最适氮源为硝酸铵、最适Ca~(2+)、Mg~(2+)浓度均为0.02%,差异有统计学意义(P0.05)。培养基优化结果显示:C源0.1%,N源0.2%,Mg~(2+)0.04%,Ca~(2+)0.04%为最佳的培养基成分组合。产酶条件筛选结果显示:培养基p H值为6、培养温度为55℃时菌株产酶水平最高,差异有统计学意义(P0.05)。结论:培养基的优化及最适的产酶条件能提高嗜热菌Anoxybacillus sp.DL4产淀粉酶能力,Ca~(2+)、Mg~(2+)离子对菌株产淀粉酶有促进作用。     

一株产黑色素海洋真菌的分离、鉴定及色素性质的研究

从海州湾筛选产黑色素的真菌,确定其分类地位,并对其所产色素的性质进行研究。对海州湾海域采集的海水、海葵、海泥等样品进行富集培养,筛选具有产色素能力的真菌。利用摇瓶发酵进行复筛,筛选得到产黑色素的菌株,对筛选得到的菌株进行鉴定,然后对其所产色素进行粗提,并对性质进行测定。通过筛选获得一株产黑色素丝状真菌菌株CXPF01,结合形态学观察和ITS分子序列分析将CXPF01鉴定为Cladosporium cladosporioides。所产黑色素在220 nm处具有最大吸收值。光照对色素的稳定性影响不显著,高温和酸碱对黑色素稳定性具有显著影响。该色素对Staphylococcus aureus和Escherichia coli具有抑制作用。该真菌所产黑色素对在光照条件下有较好的稳定性并具有抑菌作用。     

尖顶羊肚菌单孢菌株群体培养特性研究

目的:羊肚菌不同分离物在培养过程中形态学变化较大且极不稳定,通过同一子实体不同单孢菌株在不同培养基上的培养特性研究特别是产菌核能力的变化回答这些变化是否是由于多核菌株的不稳定性引起.方法:以不同来源尖顶羊肚菌单孢菌株为材料,并以粗柄羊肚菌为对照,研究了菌株在不同培养基上的培养特性,并对同一子实体及不同子实体产菌核和不产菌核单孢进行配对培养.结果:尖顶羊肚菌单孢菌株按培养特征可分为9类,同等条件下每一菌株的培养特性保持稳定;在综合马铃薯葡萄糖培养基(CPDA)、葡萄糖硝酸钠琼脂培养基(GN)、酵母膏胨葡萄糖琼脂培养基(YPD)进行转接培养时,可成功地将产菌核菌株转化为不产菌核菌株;尖顶羊肚菌同一子实体及不同子实体各产菌核单孢菌株产核数量及分布变化很大,交配后单孢之间性状会发生较大变化,包括菌核形态、菌丝形态、生长势,特别是产菌核能力会消失和发生转移.     

一株产胆固醇氧化酶的不动杆菌的筛选鉴定及特性研究

目的：筛选鉴定产胆固醇氧化酶的菌株并对其酶的性质及发酵条件进行初步的研究。方法：利用唯一碳源的胆固醇平板筛选,酶活测定比较得酶活力最高的菌株;生理生化试验结合16S rDNA序列分析鉴定其种属,单因素及正交实验优化培养基及发酵条件。结果：所得菌株H4与产不动杆菌（Acinetobacter）有最近的亲缘关系,其胆固醇氧化酶作用的最适温度和pH分别为37℃和8.0,金属离子Mg2＋、Zn2＋、Fe2＋对该酶具有一定激活作用,菌株产酶的最适培养基为（g/L）：胆固醇1.5,蔗糖5,蛋白胨7,硝酸铵3,吐温1.0,pH7.5;最适培养条件为33℃,15mL培养基/100mL三角瓶,摇床培养（200r/min）48h,优化后发酵液酶活达135.8U/L。结论：获得了1株产胆固醇氧化酶的菌株H4,并初步鉴定为不动杆菌（Acinetobacter）。     

阿拉伯胶酶产生菌株的筛选及其发酵培养基的优化

从土壤中筛选产阿拉伯胶酶的微生物菌株,并通过紫外诱变选育后得到高产突变菌株ZHB05F,依据菌落和孢子形态特征初步鉴定为镰刀茵(Fusarium sp.).通过单因素试验,优化了产酶培养基的主要组分的浓度和pH值,得到最佳的产酶发酵培养基组成为:阿拉伯胶30 g/L,(NH4)2SO4 8 g/L,K2HPO4 1g/...     

啤酒酵母产子囊抱子及获得单倍体细胞条件的研究

啤酒酵母在不同产孢培养基上表现出不同的产孢率,随菌株不同各有其最适的产孢培养基。试验表明,4号培养基对供试菌株均表现出较高的产孢率,其次是1号培养基,用2％蜗牛酶于37℃水浴酶解3小时即可除去子囊壁;将酶解液置于58℃水浴保持8分钟,可杀死营养体细胞;用1％胰蛋白酶或超声波(150W,3分钟)处理,可使50％以上的孢子分散开。     

A study of hepatic metabolism of thyroxine in BB/W rats treated with L-thyroxine

The effect of thyroxine (T4) on T4 conversion to triiodothyronine (T3) and reverse T3 (rT3) was studied in BB/W rats. A colony of 38 BB/W rats was obtained and half were treated with thyroxine (T4), 1 mg per liter of drinking water. At 106 days of age the following groups were identified: nondiabetic, no T4 treatment, 8 rats; nondiabetic, T4 treated, 8 rats; diabetic, no T4 treatment, 10 rats; diabetic, T4 treated, 7 rats. All animals with diabetes were treated with insulin. T4 conversion to T3 and rT3 was assessed in liver homogenates in 0.1 M Tris-HCl buffer, pH 7.4, with or without 5 mM dithiothreitol (DDT). Serum T4 and rT3 were significantly elevated in both T4-treated groups (P less than 0.001), while serum T3 was not affected in either. Basal T4 deiodination to T3 by the liver homogenate did not change on treatment with T4; the addition of DTT increased T3 production in the homogenate from T4 treated nondiabetic animals (P less than 0.05). In both nondiabetic and insulin-treated diabetic rats there was no effect of T4 on the rate of rT3 production. Since, in the rat, 30-40% of circulating T3 is a direct contribution of thyroid gland secretion, and that would be absent in our T4-suppressed animals, the normal serum T3 may reflect increased absolute peripheral T3 production from the greater concentration of circulating T4.     

Monocytes and platelets share the glycoproteins IIb and IIIa that are absent from both cells in Glanzmann''s thrombasthenia type I.

The possibility that thyroxine (T4) itself exerts the hormonal effect in vivo on the rat liver nuclear receptor was studied with the aid of iopanoic acid (IOP), an inhibitor of the conversion of T4 into tri-iodothyronine (T3). After administration of 2.4 micrograms of T4/100 g body weight to hypothyroid rats for 7 days, T4 and T3 concentrations in serum and in the liver nuclear non-histone protein (NHP) were all increased to the hyperthyroid range. Hepatic mitochondrial alpha-glycerophosphate dehydrogenase (alpha-GPD) activity and DNA content increased significantly. The equilibrium association constant (Ka) of the nuclear T3 receptor was unchanged and the maximal binding capacity (Cmax.) increased 1.4-fold. Simultaneous administration of IOP (5 mg/100 g body weight) to the rats given 2.4 micrograms of T4/100 g body weight completely blocked the conversion into T3. The serum T4 was even more increased, whereas the serum T3 decreased to the hypothyroid range. Although the NHP-bound T4 was at a concentration comparable with the rats given T4 alone, no NHP-bound T3 was detected. Yet the alpha-GPD activity was elevated 2.8-fold and the DNA content increased to the same extent as observed in the rats given T4 alone. The Ka and Cmax. of the nuclear receptor were significantly decreased. After administration of 48 or 480 micrograms of T4/100 g body weight for 3 days, serum T4 and T3 were markedly increased. The NHP-bound T3 was also increased, but no NHP-bound T4 was detected. The alpha-GPD activity was markedly elevated, but the DNA content was unchanged. The Cmax. per g of liver was increased, whereas the Ka remained unchanged. Simultaneous administration of IOP to these animals could not completely block the T4 conversion. The observed hormonal effects in the absence of nuclear T3 indicate that T4 possesses the intrinsic hormonal activities on the rat liver. T4 is less potent in induction of alpha-GPD activity but as potent in increment of hepatic DNA as T3. Although the binding site for T4 is not fully characterized, it appears to be acidic NHP. T4 is an active hormone, yet is also a prohormone of T3, offering the closest analogy with testosterone.     

The effect of extrathyroidal conversion inhibitor from food-deprived animals on iodothyronines deiodination by pituitary and cerebral cortical homogenates in vitro

The influence of an inhibitor of iodothyronines' extrathyroidal conversion on T4, T3 and rT3 deiodination by adult pig pituitary and cerebral cortical homogenates has been investigated. The homogenates were incubated with T4, T3 and rT3 in the presence of 5 mM dithiothreitol and evaporated diethyl ether extracts of sera obtained from fed and starved (1-14 days) rabbits. The extracts had no influence either on T4 to T3 or on T4 to rT3 conversion in cerebral cortex. Deiodination of rT3 to 3,3'-T2 in that tissue was significantly inhibited only by the extracts of sera obtained from 4 days starved rabbits. Inner-ring deiodination of both rT3 and T3 was not changed by the extracts got from short-term (1-4 days) fasted animals but was significantly reduced by the extracts from long-term (7-14 days) food-deprived subjects. Pituitary conversion of T4 to T3 was diminished by 35% in the presence of sera extracts gained from 1-9 days fasted rabbits and by about 50% on day 14 of fasting, but only the latter change was statistically significant. Short-term fasting inhibited T4 to rT3 conversion on days 2 and 4. Both deiodinations of rT3 and 5-deiodination of T3 were affected by extracts of sera collected during long-term fasting.     

T4 endocytosis and phosphorylation induced by phorbol esters but not by mitogen or HIV infection

The T4 (CD4) molecule has been shown to facilitate the interactions of T cells with HLA class II determinants, to function as a signal transducing molecule, and to serve as a receptor for HIV. Recent studies demonstrated that both phorbol esters and antigen stimulation induced the rapid and transient modulation and phosphorylation of T4 on an IL-2-dependent line of cloned peripheral blood T4+ cells. In the current study, we define the kinetics of T4 phosphorylation and internalization induced by phorbol esters and determine the extent to which this metabolic pathway is required for T cell proliferation, activation, and HIV infection. On both peripheral blood T4+ cells and the T cell line Sup-T1, the modulation and internalization of surface T4 induced by phorbol 12, 13-dibutyrate (PDB) was preceded by rapid and transient phosphorylation. On both cell types, by 48 h, T4 was reexpressed on the cell surface in a nonphosphorylated form and was shown to be resistant to phosphorylation and internalization when these cells were reexposed to PDB. In contrast, T4 on the surface of PBL was neither phosphorylated nor down-modulated when PBL were stimulated by PHA, indicating that these effects were not simply the result of T cell activation or proliferation. In additional studies, we demonstrate that this pathway for T4 phosphorylation and internalization is not required for HIV infection by showing that 1) the binding of the HIV gp 120 envelope to T4 does not induce phosphorylation of T4, 2) Sup-T1 cells that are rendered resistant to phorbol ester-induced T4 internalization and phosphorylation by prolonged culture in PDB remain highly susceptible to HIV infection, and 3) clones of HIV-producing cells expressing high levels of surface T4 that is complexed with viral envelope remain susceptible to PDB-induced modulation of T4. This observation suggests that, at least on lymphoid cells, HIV penetration does not occur exclusively by R-mediated endocytosis.     

Effect of propranolol on the metabolism of thyroid hormones

The effect of propranolol on thyroxine (T4) and 3,5',3'-triiodothyronine (T3) plasma concentrations, fractional disappearance rates (k), metabolic clearance rates (MCR) and catabolic rates has been investigated in young pigs. The animals were examined in the period 18-24 h after feeding. Plasma concentrations of T3 were lower after treatment with propranolol, but T4 was not significantly affected. The k value of T4 was decreased by propranolol but that for T3 was unaffected. The MCR of T4 and the catabolic rates of both hormones were reduced by propranolol. The reduction in metabolic rate after propranolol is thus probably related to its action on thyroid hormone metabolism.     

Host- and Phage-Mediated Repair of Radiation Damage in Bacteriophage T4

T4+ exhibits increased ultraviolet sensitivity on derivatives of Escherichia coli K12 or B lacking deoxyribonucleic acid (DNA) polymerase I. However, the sensitivity of T4v is not affected by the absence of host DNA polymerase. T4x and T4y also show increased sensitivity on DNA polymerase-deficient strains, but to a lesser extent than observed with wild-type T4. When T4x or T4y, but not T4+, are plated on a double mutant lacking both DNA polymerase and the uvrA gene product, a partial suppression of the polymerase effect is observed. Host ligase appears to be able to suppress to some extent the T4y phenotype but has no effect on wild-type T4 or other T4 mutants. T4xv incubated in E. coli B or B(s-1) in the presence of chloramphenicol (50 mug/ml) shows increased resistance over directly plated irradiated phage. Increased survival under the same conditions was not observed with T4+ or other T4 mutants. The repair of X-ray-damaged T4 was investigated by examining survival curves of T4+, T4x, T4y, T4ts43, and T4ts30. The repair processes were further defined by observing the effects of plating irradiated phage on various hosts including strains lacking DNA polymerase I or polynucleotide ligase. Two classes of effects were observed. Firstly, the x and y gene products seem to be involved in a repair system utilizing host ligase. Secondly, in the absence of host DNA polymerase, phage sensitivity is increased in an unknown manner which is enhanced by the presence of host uvrA gene product.     

Differential regulation of IFN-gamma, TNF-alpha, and IL-10 production by CD4(+) alphabetaTCR+ T cells and vdelta2(+) gammadelta T cells in response to monocytes infected with Mycobacterium tuberculosis-H37Ra.

Mycobacterium tuberculosis bacilli readily activate CD4(+) and gammadelta T cells. CD4(+) and gammadelta T cells were compared for their ability to regulate IFN-gamma, TNF-alpha, and IL-10 production, cytokines with significant roles in the immune response to M. tuberculosis. PBMC from healthy tuberculin positive donors were stimulated with live M. tuberculosis-H37Ra. CD4(+) and gammadelta T cells were purified by negative selection and tested in response to autologous monocytes infected with M. tuberculosis. Both subsets produced equal amounts of secreted IFN-gamma. However, the precursor frequency of IFN-gamma secreting gammadelta T cells was half that of CD4(+) T cells, indicating that gammadelta T cells were more efficient producers of IFN-gamma than CD4(+) T cells. TNF-alpha production was markedly enhanced by addition of CD4(+) and gammadelta T cells to M. tuberculosis infected monocytes, and TNF-alpha was produced by both T cells and monocytes. No differences in TNF-alpha enhancement were noted between CD4(+) and gammadelta T cells. IL-10 production by M. tuberculosis infected monocytes was not modulated by CD4(+) or gammadelta T cells. Thus CD4(+) and gammadelta T cells had similar roles in differential regulation of IFN-gamma, TNF-alpha, and IL-10 secretion in response to M. tuberculosis infected monocytes. However, the interaction between T cells and infected monocytes differed for each cytokine. IFN-gamma production was dependent on antigen presentation and costimulators provided by monocytes. TNF-alpha levels were increased by addition of TNF-alpha produced by T cells and IL-10 production by monocytes was not modulated by CD4(+) or gammadelta T cells.     

Amiodarone inhibits T4 to T3 conversion and alpha-glycerophosphate dehydrogenase and malic enzyme levels in rat liver

Amiodarone has been found to decrease serum T3 by blocking peripheral T4 5'-deiodinase. This reduction in T3 levels may contribute to the effectiveness of this drug in moderating cardiac arrhythmias. To further characterize the effect of amiodarone on thyroid hormone metabolism and biological action, male Sprague-Dawley rats were thyroidectomized and then fed 500 ug T4 or 50 ug T3 and 500 mg amiodarone/kg of powdered diet for 6 to 8 weeks. Hepatic and cardiac levels of T4, T3, alpha-glycerophosphate dehydrogenase (GPD) and malic enzyme (ME) were used as indicators of thyroid hormone availability and action at the cellular level. Conversion of T4 to T3 was measured in liver homogenates. Serum TSH, T4 and T3 were also measured. Amiodarone reduced hepatic GPD and ME in thyroidectomized rats receiving dietary T4. Liver T4 levels were significantly increased by amiodarone and the T3/T4 ratio was reduced (P less than .05). Amiodarone inhibited hepatic T4 to T3 conversion and decreased serum T3. The decreased T3 action at the cellular level, indicated by the reduction in hepatic GPD and ME, is not due to pharmacologic effects of amiodarone since these enzyme levels were not affected by amiodarone in thyroidectomized rats replaced with T3.     

Triiodothyronine but not thyroxine accelerates myofibrillar proteolysis via ATP production in cultured muscle cells

These experiments were done to clarify that the differential effects of thyroxine (T(4)) and triiodothyronine (T(3)) on skeletal muscle protein turnover are caused by their roles on ATP production. Primary cultured chick muscle cells were treated with a physiological level of T(4) (60 ng/ml), T(3) (12 ng/ml), or ATP (0.5 mM) for 6 days and the protein content, ATP production, proteasome activity, and myofibrillar protein breakdown were measured. The protein content measured as an index of cell growth was not affected by T(4), T(3), or ATP. The cellular ATP level was increased by T(3) and ATP, but not by T(4). Proteasome activity and N(tau)-methylhistidine (MeHis) release measured as an index of myofiblillar protein breakdown was also increased by T(3) and ATP, but not by T(4). These results indicate that T(3) but not T(4) increases ATP production followed by an increase in proteasome activity, and thus stimulates myofibrillar proteolysis.     

Selective down modulation of L3T4 molecules on murine thymocytes by the tumor promoter, phorbol 12-myristate 13-acetate

Treatment of murine thymocytes, but not mature peripheral T cells, with the tumor promoter, phorbol 12-myristate 13-acetate (PMA), 3 results in a rapid disappearance of L3T4 molecules from the surface of thymocytes. The effect of PMA on L3T4 molecules persists in vitro for at least 72 hr. Down modulation of L3T4 molecules was PMA dose-dependent and temperature-dependent. L3T4 molecules on cortisone-resistant thymocytes were significantly less sensitive to the effect of PMA than were L3T4 molecules on cortisone-sensitive thymocytes. Down modulation of L3T4 molecules on thymocytes did not interfere with their capacity to respond to concanavalin A or activation signals delivered via their T cell receptors. The difference in the ability of thymocytes and peripheral T cells to respond to PMA cannot be explained by differences in the number of PMA receptors. Both thymocytes and peripheral T cells have PMA receptors in the range of 1 to 1.5 X 10(5) receptors/cell. However, there is a small difference in the affinity (Kd) of the receptors on thymocytes (Kd = 30 to 40 nM) and peripheral T cells (Kd = 10 to 15 nM). Immunofluorescent staining revealed that the down modulation of L3T4 molecules by PMA was a result of internalization of L3T4 molecules. After down modulation, L3T4 could be readily detected on the cytoplasm of thymocytes. These findings suggest that L3T4 molecules on thymocytes may be subject to different regulatory signals than L3T4 molecules on peripheral T cells.