Organic Chemical Investigation of Novel Biologically Active Compounds, Syntheses and Reactions, Evaluation of Biological Activities.

(1)在相模子、茨城和京都采集了雌性独居黄蜂标本。提取冻干后的蛇毒囊，反复进行反相高效液相色谱法，同时用龙虾行走腿神经肌肉制剂检测神经活动，得到纯化的蛇毒。分离出的蛇毒主要通过质谱学、MALDI-TOF-CID法和/或梯形序列法进行结构鉴定，最后通过基于Fmoc-L-氨基酸策略的固相合成法确定其结构。Eumenine mastoparan-AF是从Eumenine黄蜂的毒液中分离得到的。黄缘Anterhynchium visomarinata Micado.其结构和神经活性均与大黄蜂的马蜂胶相似。新的和独有的神经毒素--α-和β-松材线虫毒素是从龙虾、萨马里亚目和斑潜蝇的毒液中分离得到的，这些毒素通过p-…增强龙虾腿肌的突触传递。(2)合成了2-甲基和4-甲基-1，25-二羟基维生素D_3的所有8个可能的A环非对映异构体，以考察A环构象[α-型(活性形式；lα-OH为赤道)]与生物活性的关系(以亲和力为指标)。合成了2-甲基-20-表-L、25-二羟基维生素D3等8种可能的A环非对映异构体。2-甲基和4-甲基的引入在一定程度上会导致平衡向任一方向移动。从3-羟基-2-甲基丙酸甲酯的(S)或(R)-异构体出发，得到了总收率较高的A环部分(烯炔类)，然后通过Trost程序与CD-环侧链部分偶联得到了2-甲基类似物。类似地，从1，3-丙二醇开始，4-甲基类似物的A环部分被制备为外消旋形式，分离后，经过相同的步骤得到4-甲基立体异构体。在用牛胸腺进行的VDR亲和中，2-甲基异构体的效力高度依赖于A环取代基的立体化学。因此，2α-Me-1α-OH，3β异构体的亲和力比天然α，25-(OH)_2-D_3(归一化，以此类推)高4倍。该异构体在动员钙离子的能力上也提高了4倍，在HL-60细胞分化中的能力提高了2倍。此外，A环和侧链的双重修饰的立体异构体2α-Me-20-epi-1α，25-(OH)_2-D_3对VDR的亲和力提高了12倍，钙动员能力提高了7倍，对HL-60细胞的分化能力提高了590倍，是迄今报道的最有效的类似物。与2-甲基系列相比，4-甲基-1，25-(OH)_2-D_3的所有非对映异构体的VDR亲和力均小于10^21-10^3。从核磁共振的构象分析来看，A环中2α-Me-1α，25-(OH)_2-D_3的构象以椅型为主，而2β-Me异构体则以半船型为主。进一步的合成(2-位各种取代基，构象受限)和生物学(细胞凋亡，转录活性)的研究正在进行中。较少

(1) The specimens of female solitary wasps were collected in Sagamiko, Ibaraki, and Kyoto. The lyophilized venom sacs were extracted and subjected repeatedly to reverse-phase HPLC, accompanied by testing neuro activity using neuromuscular preparations of walking leg of lobster, to give purified venoms. The structures of the isolated venoms were estimated mainly by the leading tequniques of mass spectrometry, MALDI-TOF-CID method andlor Ladder sequence method, and finally determined by solid phase synthesis based on the Fmoc- L-amino acid strategy. Eumenine mastoparan-AF was isolated from the venom of the Eumenine wasp. Anterhynchium flavomarginatum micado. Both the structure and theneuroactivity were similar to those ofmastoparan of a hornet wasp. Novel and uniqueneurotoxins, alpha- and beta-pompilidotoxins were isolated from the venoms of the Pompilidaewasps, Anoplius samariensis and Batozonellus maculifrons, These venoms potentiatessynaptic transmission of lobster leg muscle by the p … More resynaptic mechanisms.(2) All eight possible A-ring diastereomers of both 2-methyl- and 4-methyl- 1,25-dihydroxyvitamin D_3 were synthesized to investigate the relationship of the A-ring conformation [alpha-form<double arrow>beta-form (active form ; lalpha-OH is equatorial)] to the biological activities C/DR affinity as an index). Also, all eight possible A- ring diastereomers of 2-methyl-20-epi-l, 25-dihydroxyvitamin D3 were synthesized. Introduction of 2-methyl- and 4-methyl-group would result in equibrium shift to either one way in certain extent. Starting from either (S)- or (R)-isomer of methyl 3-hydroxy-2-methyl-propionate, the A-ring portions(enynes) were obtained in good overall yields, which were subsequently coupled with the CD-ring-side chain portion by the Trost' Procedure to give the 2-methyl analogues. Similarly, starting from 1,3-propanediol, the A-ring portions of 4-methyl analogues were prepared as racemic forms, which, after separation, were subjected to the same procedure to give 4-methyl steroisomers. In VDR affinity using bovine thymus, the potency of 2-methyl isomers was highly dependent on the stereo-chemistry of the A-ring substituents. Thus, 2alpha-Me-1alpha-OH,3betaisomer showed 4-fold higher affinity than natural lalpha, 25-(OH)_2-D_3 (normalized, and so forth). This isomer also exhibited 4-fold higher potency in Ca mobilization, and 2-fold higher potency in HL-60 cell differentiation. Furthermore, double modified steroisomer of A-ring and side chain, 2alpha-Me-20-epi-1alpha, 25-(OH) _2-D_3 showed 12 fold higher in VDR affinity, 7 fold higher in Ca mobilization and 590 times higher potency in HL-60 cell differentiation, which is the most potent analogue reported to date. In contrast to 2-methyl series, VDR affinities of all diastereomers of 4-methyl-1,25-(OH) _2-D_3 were less than 10^21-10^3. From the conformational analysis by NMR spectroscopy, chair-wform is predominant in A-ring coconformation of 2alpha-Me-1alpha, 25-(OH) _2-D_3, while 2beta-Me isomer prefer half-boat-beta-form in A-ring. Further synthetic (various substituents in 2-position, conformational restricted) and biological (apotosis, transcriptional potency) studies are in progress. Less