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

(1)分别在相模美子、茨城县和京都采集雌独居胡蜂标本。提取冻干毒液囊，反复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性，得到纯化毒液。主要采用质谱法、MALDI-TOF-CID法和阶梯序列法对分离毒液的结构进行鉴定，最后采用基于Fmoc- l -氨基酸策略的固相合成方法对分离毒液的结构进行鉴定。从蜂蜂的毒液中分离得到蜂蜂乳突蛋白af。黄精凤仙花。其结构和神经活性均与大黄蜂的mastoparan相似。从pompilidaewasp、anplius samariensis和Batozonellus maculifrons的毒液中分离出了新的和独特的神经毒素，α -和β -pompilidotoxins，这些毒液通过p…More突触机制增强了龙虾腿肌肉的突触传递。(2)合成了2-甲基-和4-甲基- 1,25-二羟基维生素D_3的8种可能的a环非对映体，研究了a环构象[α -型<双箭头> β -型（活性型；α - oh为equatorial）]与生物活性C/DR亲和度的关系。同时，合成了2-甲基-20-epi- 1,25 -二羟基维生素D3的8种可能的A-环非对映体。2-甲基和4-甲基的引入会在一定程度上导致平衡向任何一个方向转移。从3-羟基-2-甲基丙酸甲酯的(S)-或(R)-异构体开始，以良好的总产率得到a环部分（烯），随后通过Trost过程将其与cd环侧链部分偶联得到2-甲基类似物。同样，从1,3-丙二醇开始，4-甲基类似物的a环部分以外消旋形式制备，分离后，经过相同的步骤得到4-甲基甾体异构体。在牛胸腺的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，2 α - me -1 α -OH，3 β异构体的亲和力比天然的α， 25-(OH)_2-D_3（归一化，依此类推）高4倍。该异构体对Ca的动员作用和HL-60细胞的分化作用也分别提高了4倍和2倍。此外，双修饰的a环和侧链体异构体2alpha-Me-20-epi-1alpha， 25-(OH) _2-D_3对VDR的亲和力提高了12倍，对Ca的动员能力提高了7倍，对HL-60细胞分化的效力提高了590倍，是迄今为止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的所有非对映体的VDR亲和都小于10^21-10^3。从核磁共振构象分析来看，2 α - me -1 α, 25-(OH) _2-D_3在a环构象中以椅形为主，而2 β - me在a环构象中以半船形为主。进一步的合成（各种2位取代基，构象受限）和生物学（细胞凋亡，转录效力）研究正在进行中。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