绞股蓝总皂甙含量与其营养器官和生长期相关性的研究

应用组织化学和植物化学方法,研究绞股蓝总皂甙含量与其营养器官和生长期的相关性。结果表明,不同营养器官中绞股蓝总皂甙含量叶最高,茎次之,根茎和根最低。绞股蓝地上部分总皂甙含量从营养生长→开花→结果不同生长期呈现由低→高→低的变化趋势,其中7、8、9月的开花期绞股蓝总皂甙含量达到顶峰,为药材最佳采收时间。研究还得出,绞股蓝叶的形态结构特征与其总皂甙含量相关,可作为选育绞股蓝优良品种的指标。     

绞股蓝龙须茶的人参皂甙组织定位及皂甙含量的季节变化

应用组织化学定位技术及植物化学方法,研究了人参皂甙在商品绞股蓝龙须茶中的组织分布状态及绞股蓝总皂甙在不同季节中的含量变化。结果表明,绞股蓝内的人参皂甙遇特定的显色剂出现深浅不同的红色,色度与人参皂甙含量呈正相关。绞股蓝叶内的人参皂甙主要分布在叶肉组织,茎内的人参皂甙主要分布于维管束的韧皮部薄壁细胞中,同化组织、厚角组织和表皮也有少量分布．木质部和髓薄壁组织中无皂甙分布。在4月至11月的龙须茶采收期中,总皂甙含量呈逐渐上升的势态,4月最低．9月最高,含量变化与月总降雨量呈显著的正相关。研究认为在8月后湿润凉爽气候条件下采摘绞股蓝芽尖加工而成的龙须茶品质最好。     

金花茶鲜叶的杀青技术研究

以金花茶(Camellia nitidissima C.W.Chi)鲜叶为原料,以金花茶叶中主要活性成分叶绿素、茶多酚、黄酮的含量为指标,比较和分析了不同方式(漂烫、蒸汽、微波)对金花茶鲜叶杀青影响,以获得金花茶叶的最佳杀青方式及杀青条件,为金花茶资源的充分开发利用提供参考。研究结果表明:漂烫杀青对金花茶鲜的叶绿素和茶多酚的破坏非常大,不利于活性成分的保留;蒸汽杀青对金花茶鲜叶中黄酮的含量影响较大,与其它2种杀青方式相比含量最低;而采用微波杀青的金花茶叶中各活性成分含量均为最高。其中微波火力为528 W杀青时间90 s处理的金花茶鲜叶的叶绿素含量为0.881 04 mg/g,茶多酚含量为6.9%,黄酮含量为7.12%,可作为金花茶鲜叶的最佳杀青条件。     

金花茶鲜叶微波杀青工艺技术研究

以金花茶鲜叶为原料,以鲜叶微波杀青后茶多酚含量、黄酮溶出量、叶绿素含量作为评价指标,先对杀青时间、投叶层数、微波火力3个因素进行单因素水平实验;并在此基础上进行L~9(3~3)正交试验;最后运用方差、极差分析确定最佳的金花茶鲜叶微波杀青条件,建立了金花茶鲜叶保质高效杀青工艺。研究结果表明:金花茶鲜叶微波杀青时,投叶层数为6层、微波功率528 W及微波杀青时间为90 s时金花茶叶中叶绿素含量为0.612 mg·g~(-1),茶多酚含量为8.797%,黄酮溶出量为9.217%,内质成分保持最优。本研究结果为金花茶鲜叶的开发利用提供了技术支持。     

绞股蓝总皂甙提取方法的比较研究

目的：探讨绞股蓝总皂甙最佳的提取方法。方法：首先选用水、80％甲醇、75％乙醇和氯仿等作为溶剂提取绞股蓝总皂甙,从中选择出2种较优试剂,再以回流提取法、索氏提取法、超声波法等提取方法对绞股蓝的总皂甙进行提取,从而选出一种优良的提取方法。并且采用分光光度法测定了绞股蓝总皂甙的含量。结果：研究表明,75％乙醇为绞股蓝总皂甙提取的最佳溶剂,乙醇回流提取法为最优方法。结论：以75％乙醇为提取溶剂的回流提取法是一种高效、简便、节能的绞股蓝总皂甙的提取方法。     

绞股蓝营养器官各发育阶段结构与总皂甙含量相关性的研究

应用组织化学定位技术及植物化学方法,研究了不同发育阶段绞股蓝茎叶的结构与其中人参皂甙分布积累和总皂甙含量的关系。绞股蓝内的人参皂甙通过特定的显色剂能显示从紫红到浅红的颜色反应,颜色深浅与人参皂甙含量呈正相关。结果表明,叶内的皂甙主要分布在叶肉组织内,未分化的叶肉组织比成熟组织含量高,衰老的叶片不含皂甙。茎内的皂甙主要分布于维管束的外生韧皮部薄壁细胞中,表皮、同化组织、厚角组织及周皮的栓内层也有少量分布,木质部和髓薄壁组织无皂甙积累;幼茎所含总皂甙高于发育成熟的茎和老茎,含量最高的则是具有贮藏功能的根状茎。在绞股蓝全草中,叶的总皂甙含量高于相应发育阶段的茎,而以当年生根状茎的含量最高。提出了皂甙在叶内积累、茎内运输和地下茎内贮藏的观点及科学采收绞股蓝的建议。     

乌龙茶鲜叶游离氨基酸组成轮廓的模式识别研究

为探明乌龙茶品种鲜叶游离氨基酸的组成轮廓特征，以绿茶品种春季幼嫩鲜叶(一芽二或三叶)和成熟鲜叶(中小开面二至四叶)为对照，对采自乌龙茶品种春、秋两季的幼嫩和成熟鲜叶进行高效液相色谱检测分析和统计模式识别。结果表明,茶树品种、芽叶嫩度和采摘季节对供试鲜叶游离氨基酸组成轮廓均有影响。全部供试茶样可按茶树品种特性划分成乌龙茶与绿茶品种2大类群，且芽叶嫩度对乌龙茶鲜叶游离氨基酸组成轮廓的影响仅次于茶树品种特性引起的茶样组群差异。绿茶与乌龙茶品种春季幼嫩鲜叶、绿茶与乌龙茶品种春季成熟鲜叶、春季乌龙茶品种幼嫩与成熟鲜叶、秋季乌龙茶品种幼嫩与成熟鲜叶、春季和秋季乌龙茶品种幼嫩鲜叶及春季和秋季乌龙茶品种成熟鲜叶间存在客观的类群区分。不同鲜叶来源对供试茶样的游离氨基酸总量及大部分游离氨基酸组分均有显著或极显著影响。较高的甘氨酸、亮氨酸、赖氨酸含量和较低的γ-氨基丁酸含量是乌龙茶品种有别于绿茶品种春季鲜叶的主要差异性成分；相同采摘季节的乌龙茶品种幼嫩鲜叶相较成熟鲜叶拥有更高的精氨酸含量，而不同芽叶嫩度的春季和秋季乌龙茶鲜叶存在不同的差异特征组分。因此，茶树品种遗传特性是影响茶树鲜叶游离氨基酸组成轮廓的主导...     

绞股蓝总皂甙对实验性高脂血症大鼠血脂和脂质过氧化水平的影响

目的：研究绞股蓝总皂甙对实验性高脂血症大鼠血脂的调节机制并探索其抗脂质过氧化的机理。方法：用高脂饲料喂饲大鼠6周,复制成高脂血症模型,而后进行4周高、低两种剂量的绞股蓝和阳性对照药血脂康实验性治疗。给药4周后处死动物,比较血清总胆固醇及甘油三酯含量、血清高密度脂蛋白胆固醇和低密度脂蛋白胆固醇含量,同时测定血清SOD、CAT和MDA的浓度。结果：高、低剂量的绞股蓝总皂甙均可显著降低大鼠血清低密度脂蛋白胆固醇、总胆固醇、甘油三脂浓度,提高高密度脂蛋白胆固醇水平 绞股蓝总皂甙组大鼠血清超氧化物歧化酶、谷胱甘肽过氧化物酶及过氧化氢酶显著升高,脂质过氧化产物水平显著降低。结论：绞股蓝总皂甙可治疗高血脂症大鼠血脂异常并有较明显的抗脂质过氧化作用。     

咖啡叶茶的研制

以咖啡鲜叶为原料,采用晒青绿茶的加工工艺,成功研制出咖啡叶子茶,该茶条索肥硕、汤色黄绿明亮、滋味醇厚并伴有淡淡花果香。其加工工艺为:咖啡鲜叶→摊晾(摊晾厚度在15-20厘米)→杀青(杀青的锅温在240℃-300℃之间)→揉捻→晒干。     

绞股蓝人参皂甙的组织化学定位及其含量的变化

应用光镜技术、组织化学定位及植物化学方法,研究了人参皂甙在绞股蓝营养器官中的积累分布状态以及不同生长期、不同器官、不同性别之间的绞股蓝总皂甙含量的动态变化。结果表明,绞股蓝人参皂甙主要分布在营养器官的同化组织及韧皮部薄壁细胞中,厚角组织、表皮及周皮的栓内层也有少量分布,木质部和髓薄壁组织中无皂甙分布;叶中皂甙积累最多,茎次之,根最少。绞股蓝在营养生长期→花果期→枯萎期的生长发育过程中,其地上部分的皂甙含量呈现出低→高→低的变化趋势;叶的含量高于茎,雄株的含量高于雌株。从而认为在9-10月的花果期采收绞股蓝的地上部分而保留地下茎和根,有利于药材品质和产量的提高,又有利于药用资源的可持续开发利用。     

Inhibition of stimulation in murine mixed lymphocyte cultures with an alloantiserum directed against a shared Ia determinant.

Pools of high titered alloantisera were raised by immunizing (B10.A/SgSn X A/WySn)F1 mice with C57BL/10Sn(B10) spleen cells. This serum (F1 anti-B10), when added to one-way mixed lymphocyte cultures (MLC), inhibited stimulation of B10.A splenic responders by both B10 and B10.D2/nSn irradiated, splenic stimulators. The B10 stimulation was suppressed approximately 85% whereas the mean suppression of B10.D2 stimulation was approximately 60%. In the ofrmer case, the serum contained antibodies reactive with multiple major histocompatibility complex determinants on the stimulator cells. In the latter case, the cytoxic reactivity of the serum was directed principally against an I region-associated determinant Ia.8) shared by B10 and B10.D2 and coded for by a gene(s) in the I-A subregion. The magnitude of the suppression of the response to B10.D2 cells (60%) was similar to the reduction in stimulation observed when the Ia.8 difference was eliminated genetically by using (B10 X B10.A)F1 responder cells against irradiated B10.D2 stimulators. Ihhibition of MLC by this antiserum was a function of reactivity with stimulator and not responder cells. Although some pools of F1 anti-B10 antiserum produced partial inhibition of the responder cell in a B10.D2 vs B10.Ax MLC combination, the results were inconsistent and not correlated with the anti-Ia.8 cytotoxicity titers. In addition, an F1 anti-B10 antiserum pool, which consistently failed to inhibit the responder cell, nevertheless inhibited both irradiated B10.D2 and (B10.A X B10.D2)F1 cells from stimulating B10.A responder cells. However, this same antiserum did not inhibit stimulation of B10.D2 responder cells by the (B10.A X B10.D2)F1 stimulators. Thus, the binding of antibodies to the non-stimulating antigens on the F1 stimulator cell did not interfere with the capacity of the appropriate stimulating antigens to cause stimulation. All of these results are consistent with the hypothesis that Ia allo-antigens are the major stimulating determinants of I region-associated MLC reactions.     

Matching unrelated stimuli with same discriminative functions: training order effects

Previous research has shown that after training simple discriminations (A1+/A2−, B1+/B2−), bringing these tasks under conditional control (J1–A1, J2–A2) leads to transfer of discriminative control (J1+/J2−) and to generalized matching on the basis of same discriminative functions (e.g. J1–B1, J2–B2). The same occurs when conditional discriminations are trained (D1–E1, D2–E2; F1–G1, F2–G2). When the subjects are then trained to demonstrate correct relations (D1–E1, D2–E2) when given X1 and to demonstrate incorrect relations when given X2 (XD–E), transfer of discriminative control (X1+/X2−) and generalized matching on the basis of same discriminative functions emerges (e.g. X1F1–G1, X2F1–G2). The present study investigated if these performances are dependent on the training and/or testing order. In Experiment 1, the lower-order contingency tasks were trained before the higher-order contingency tasks (A1+/A2−, B1+/B2− before J–A, and D–E, F–G before XD–E). Half the subjects received the J–B test before the more complex XF–G test (Condition A), while for the other subjects, this testing order was reversed (Condition B). Finally, all subjects received additional tests in which they were given the opportunity to demonstrate the discriminative properties of the J and X stimuli (J1+/J2−, X1+/X2−), and to match the A, J, and X stimuli with newly introduced stimuli of same discriminative properties (e.g. J1-POLITE, J2-RUDE). Experiment 2 was the same except that the training order was reversed (J–A before A1+/A2−, B1+/B2−, and XD–E before D–E, F–G). The results were affected by the training order but not by the testing order. Transfer of discriminative functions and generalized matching on the basis of same functions only occurred reliably when the lower-order contingency tasks were trained first. A stimulus-control account of the data is offered.     

Karyotype of chironomid Xenochironomus xenolabis (diptera,chironomidae)--obligatory miner of fresh water sponges

The karyotype of chironomid Xenochironomus xenolabis has been first described. Chromosomes of X. xenolabis have a peculiar spatial organization: subcentromere areas of all chromosomes join together in one common chromocenter. Seven chromosome arms (A = B = C > D > E > F > G) are conventionally recognized in the karyotype. Each arm is subdivided into sections, beginning with the centromere area. The nucleoli are located in arm B and probably D. No chromosome reorganization was found in X. xenolabis. The strong conservatism and ordered organization of the chromosome set in X. xenolabis seem to be determined by a specific life style of its larvae and, consequently, by a relatively constant habitation conditions.     

Cytochrome b561 protein family: expanding roles and versatile transmembrane electron transfer abilities as predicted by a new classification system and protein sequence motif analyses

Cytochrome b561 family was characterized by the presence of "b561 core domain" that forms a transmembrane four helix bundle containing four totally conserved His residues, which might coordinate two heme b groups. We conducted BLAST and PSI-BLAST searches to obtain insights on structure and functions of this protein family. Analyses with CLUSTAL W on b561 sequences from various organisms showed that the members could be classified into 7 subfamilies based on characteristic motifs; groups A (animals/neuroendocrine), B (plants), C (insects), D (fungi), E (animals/TSF), F (plants+DoH), and G (SDR2). In group A, both motif 1, {FN(X)HP(X)2M(X)2G(X)5G(X)ALLVYR}, and motif 2, {YSLHSW(X)G}, were identified. These two motifs were also conserved in group B. There was no significant features characteristic to groups C and D. A modified version of motif 1, {LFSWHP(X)2M(X)3F(X)3M(X)EAIL(X)SP(X)2SS}, was found in group E with a high degree of conservation. Both motif 3, {DP(X)WFY(L)H(X)3Q}, and motif 4, {K(X)R(X)YWN(X)YHH(X)2G(R/Y)} ,were found in group F at different regions from those of motifs 1 and 2. The "DoH" domain common to the NH2-terminal region of dopamine beta-hydroxylase was found to form fusion proteins with the b561 core domains in groups F and G. Based on these results, we proposed a hypothesis regarding structures and functions of the 7 subfamilies of cytochrome b561.     

Complex Genotype Mixtures Analyzed by Deep Sequencing in Two Different Regions of Hepatitis B Virus

This study assesses the presence and outcome of genotype mixtures in the polymerase/surface and X/preCore regions of the HBV genome in patients with chronic hepatitis B virus (HBV) infection. Thirty samples from ten chronic hepatitis B patients were included. The polymerase/surface and X/preCore regions were analyzed by deep sequencing (UDPS) in the first available sample at diagnosis, a pre-treatment sample, and a sample while under treatment. HBV genotype was determined by phylogenesis. Quasispecies complexity was evaluated by mutation frequency and nucleotide diversity. The polymerase/surface and X/preCore regions were validated for genotyping from 113 GenBank reference sequences. UDPS yielded a median of 10,960 sequences per sample (IQR 16,645) in the polymerase/surface region and 11_,595 sequences per sample (IQR 14,682) in X/preCore. Genotype mixtures were more common in X/preCore (90%) than in polymerase/surface (30%) (p<0.001). On X/preCore genotyping, all samples were genotype A, whereas polymerase/surface yielded genotypes A (80%), D (16.7%), and F (3.3%) (p = 0.036). Genotype changes in polymerase/surface were observed in four patients during natural quasispecies dynamics and in two patients during treatment. There were no genotype changes in X/preCore. Quasispecies complexity was higher in X/preCore than in polymerase/surface (p = 0.004).The results provide evidence of genotype mixtures and differential genotype proportions in the polymerase/surface and X/preCore regions. The genotype dynamics in HBV infection and the different patterns of quasispecies complexity in the HBV genome suggest a new paradigm for HBV genotype classification.     

Toxoid of Clostridium botulinum type F: purification and immunogenicity studies.

Toxin from Clostridium botulinum type F was recovered from dialysis cultures and partially purifed by: (i) ammonium sulfate and ethanol precipitation; (ii) O-(diethylaminoethyl)-cellulose chromatography; or (iii) diethylaminoethyl-cellulose chromatography followed by O-(carboxymethyl)-cellulose chromatography. Toxin purities as reflected by specific activity were 1.83 X 10(6), 9.8 X 10(6), and 2.0 X 10(7) mouse 50% lethal doses (LD50)/mg of N, respectively, for toxins purified by the three methods. The toxins were converted to toxoids by incubation at 35 C in the presence of 0.3 to 0.45% formalin for 21 to 35 days. Toxoids were immunogenic in guinea pigs, as demonstrated by serum antitoxin response and the immunized animals' resistance to challenge by type F botulinal toxin. The immune response to type F toxoids was lower when toxoids of serotypes A, B, C, D, and E were combined with the type F toxoid than when the type F toxoid only was administered. The toxoid prepared from the most highly purified toxin (method [iii]) conferred the highest immunity in guinea pigs at a given dose level. A relation between serum antitoxin level and resistance to challenge was observed. At least 50% of the groups of guinea pigs with 0.015 antitoxin units or more per ml survived challenge by 10(5) mouse LD50 of type F botulinal toxin. A dose of 3.75 mug of N of the most highly purified type F toxoid in combination with the other five serotypes of botulinal toxoid invoked an immune response in guinea pigs comparable to that considered adequate for the other toxoids.     

A new erythrocytic antigen of C57BL/10 (B10) mice

A new antigen, detectable on murine erythrocytes by hemagglutination assay with a (BALB/cCrl X SWR/J)F1 anti-B10.D2n/Sn alloantiserum, is described. Among the inbred and congenic mouse strains tested for reactivity with the antiserum, only the immunizing strain, B10.D2, and its congenic resistant partner, C57BL/10 (B10), reacted. Three other C57 strains, C57BL/6J, C57BL/6By, and C57L, were negative for the antigen. F1 hybrids between B10 and BALB/c, an antigen-negative strain, were positive for the antigen indicating that its expression is dominant. Typing of 39 (BALB/c X (BALB/c X B10)F1) and 62 [BALB/c X B10)F1 X BALB/c) backcross mice revealed that a single gene controls expression of the antigen. The gene is autosomal and not linked to H-2, Ly-4, or the c (albino) or b coat color genes.     

A kinetic assessment of the sequence of electron transfer from F(X) to F(A) and further to F(B) in photosystem I: the value of the equilibrium constant between F(X) and F(A)

The x-ray structure analysis of photosystem I (PS I) crystals at 4-A resolution (Schubert et al., 1997, J. Mol. Biol. 272:741-769) has revealed the distances between the three iron-sulfur clusters, labeled F(X), F(1), and F(2), which function on the acceptor side of PS I. There is a general consensus concerning the assignment of the F(X) cluster, which is bound to the PsaA and PsaB polypeptides that constitute the PS I core heterodimer. However, the correspondence between the acceptors labeled F(1) and F(2) on the electron density map and the F(A) and F(B) clusters defined by electron paramagnetic resonance (EPR) spectroscopy remains controversial. Two recent studies (Diaz-Quintana et al., 1998, Biochemistry. 37:3429-3439;, Vassiliev et al., 1998, Biophys. J. 74:2029-2035) provided evidence that F(A) is the cluster proximal to F(X), and F(B) is the cluster that donates electrons to ferredoxin. In this work, we provide a kinetic argument to support this assignment by estimating the rates of electron transfer between the iron-sulfur clusters F(X), F(A), and F(B). The experimentally determined kinetics of P700(+) dark relaxation in PS I complexes (both F(A) and F(B) are present), HgCl(2)-treated PS I complexes (devoid of F(B)), and P700-F(X) cores (devoid of both F(A) and F(B)) from Synechococcus sp. PCC 6301 are compared with the expected dependencies on the rate of electron transfer, based on the x-ray distances between the cofactors. The analysis, which takes into consideration the asymmetrical position of iron-sulfur clusters F(1) and F(2) relative to F(X), supports the F(X) --> F(A) --> F(B) --> Fd sequence of electron transfer on the acceptor side of PS I. Based on this sequence of electron transfer and on the observed kinetics of P700(+) reduction and F(X)(-) oxidation, we estimate the equilibrium constant of electron transfer between F(X) and F(A) at room temperature to be approximately 47. The value of this equilibrium constant is discussed in the context of the midpoint potentials of F(X) and F(A), as determined by low-temperature EPR spectroscopy.     

Assembly of photosystem I. II. Rubredoxin is required for the in vivo assembly of F(X) in Synechococcus sp. PCC 7002 as shown by optical and EPR spectroscopy

The rubA gene was insertionally inactivated in Synechococcus sp. PCC 7002, and the properties of photosystem I complexes were characterized spectroscopically. X-band EPR spectroscopy at low temperature shows that the three terminal iron-sulfur clusters, F(X), F(A), and F(B), are missing in whole cells, thylakoids, and photosystem (PS) I complexes of the rubA mutant. The flash-induced decay kinetics of both P700(+) in the visible and A(1)- in the near-UV show that charge recombination occurs between P700(+) and A(1)- in both thylakoids and PS I complexes. The spin-polarized EPR signal at room temperature from PS I complexes also indicates that forward electron transfer does not occur beyond A(1). In agreement, the spin-polarized X-band EPR spectrum of P700(+) A(1)- at low temperature shows that an electron cycle between A(1)- and P700(+) occurs in a much larger fraction of PS I complexes than in the wild-type, wherein a relatively large fraction of the electrons promoted are irreversibly transferred to [F(A)/F(B)]. The electron spin polarization pattern shows that the orientation of phylloquinone in the PS I complexes is identical to that of the wild type, and out-of-phase, spin-echo modulation spectroscopy shows the same P700(+) to A(1)- center-to-center distance in photosystem I complexes of wild type and the rubA mutant. In contrast to the loss of F(X), F(B), and F(A), the Rieske iron-sulfur protein and the non-heme iron in photosystem II are intact. It is proposed that rubredoxin is specifically required for the assembly of the F(X) iron-sulfur cluster but that F(X) is not required for the biosynthesis of trimeric P700-A(1) cores. Since the PsaC protein requires the presence of F(X) for binding, the absence of F(A) and F(B) may be an indirect result of the absence of F(X).