Biochemical Studies on Halocynine, a New Betaine Isolated from the Muscle of an Ascidian Halocynthia roretzi.

本研究建立了一种从海鞘Halocynthia roretzi肌肉中分离得到的甜菜碱--卤花宁的高效液相色谱测定方法，并对其在海鞘及其近缘种组织中的分布进行了研究。H.用示踪技术对roretzi进行了研究。研究结果如下：1.用紫外吸收的对二溴苯乙酮对卤代花菁进行酯化，用Shim-pack FLC-CN色谱柱，含5%CH_3CN（pH2.0）的5mMKH_2PO_4缓冲液，在262 nm处进行紫外检测。对溴苯甲酰甲基卤代花菁酯与海鞘肌肉中丰富的甘氨酸甜菜碱酯及其它紫外吸收物质分离良好。在300 nmol范围内，盐花宁酯的峰面积与盐花宁的含量呈线性关系.每100 g肌肉中可检测到50 mg以上的卤霉素。罗雷茨全年都有，但含量冬季高，夏季低。在肝脏和鳃中发现了少量的卤霉素，但在生殖腺和图尼卡中没有发现。在皱瘤海鞘的肌肉、肝脏、生殖腺和鳃中，以及在紫葡萄藻、拟花海鞘和玻璃海鞘的全身中，均检测到了从微量到30 mg/100 g的卤代半胱氨酸.将~（13 <14>C）-赖氨酸注入H. roretzi，将动物在海水箱中保持10天。然后，从肌肉中制备苦味酸提取物，并进行离子交换柱层析以分离显示放射性的卤花青。在四种不同溶剂系统的TLC上，放射性随分离的卤花青斑点迁移。这些结果清楚地表明，在H. roretzi。

The present studies were conducted to develop an HPLC method for determination of halocynine, a new betaine isolated from the muscle of an ascidian Halocynthia roretzi, and to survey its distribution in the tissues of the ascidian and in the related species. Biosynthesis of halocynine in H. roretzi was also investigated with tracer technics. Results obtained were summarized as follows.1. The authentic halocynine was esterified with UV-absorbing ,p-dibromoacetophenone and subjected to an HPLC using Shim-pack FLC-CN column and 5 mM KH_2PO_4 buffer containing 5 % CH_3CN (pH 2.0) with an ultraviolet detector operater at 262 nm. Halocynine p-bromophenacyl ester was well separated from the ester of glycinbetaine and other UV-absorbing substances which were abundant in the ascidian muscle. The linear relationship was obtained between the peak areas of halocynine ester and the amounts of halocynine up to 300 nmol.2. More than 50 mg of halocynine per 100 g was detected in the muscle of H. roretzi throughout the year, but its content was high in winter and low in summer. Small amounts of halocynine were found in the liver and gills but not in the genital gland and tunica. Halocynine ranging from a trace amount to 30 mg per 100 g was also detected in the muscle, liver, gonad and gills of Styela plicata and in the whole bodies of Botrylloides violaceus, Pseudoistoma antinboja and Ciona intestinalis.3. After ^<14>C-lysine was injected into the muscle of H. roretzi, the animals were kept in a seawater tank for 10 days. Then, a picric acid extract was prepared from the muscle and subjected to an ion exchange column chromatography to isolate halocynine, which showed a radioactivity. The radioactivity migrated with the spots of isolated halocynine on TLC with four different solvent systems. These results clearly show that halocynine is biosynthesized from lysine in H. roretzi.