Fund-Nutritional Properties of Physiological Functions of Vitamin Biz and Antioxidative Substances of Algae

采用莱希曼乳杆菌ATCC 7830微生物法和内在因子化学发光法测定了干青菜（Enteromorpha sp.）和紫菜（Porphyra sp.）的维生素B_<12>浓度。干绿紫菜和干紫紫菜的微生物测定值（68.58±2.9和32.26±1.61 μg/100g干重）与化学发光法测定值（60.20±2.21和25.07±0.54 <μg/100g干重）相同。测定了一种藻类保健食品——螺旋藻（spirulina sp.）片中维生素B_<12>的浓度。螺旋藻片的微生物测定值比化学发光法测定值高6 ~ 9倍。用全自动化学发光分析仪测定了食品中维生素B_<12>的含量，并与微生物法进行了比较。在贝类和螺旋藻中，微生物荧光法测定的值比化学发光法测定的值高6-8倍，而在其他食品中，两种方法测定的值有很好的相似性。为了明确紫菜中维生素B_<12>的生物利用度，测定了紫菜中维生素B_<12>和维生素B_<12>类似物的含量，并研究了紫菜对维生素B_<12>缺乏大鼠的影响。紫菜含有5种具有生物活性的维生素B_<12>化合物（氰基、羟基、亚硫酸盐、腺苷基和甲基钴胺素），其中维生素B_<12>辅酶（腺苷基和乙基钴胺素）约占维生素B_<12>总量的60%。9周龄维生素缺陷大鼠尿中甲基丙二酸排泄量为71.7(SE 20.2) μ mol/d，饲喂紫紫菜（10μg/kg） 20 d后，尿中甲基丙二酸排泄量（作为维生素B_<12>缺乏的指标）无法检测，肝脏维生素B_<12>（特别是腺苷钴胺素）水平显著升高。上述结果表明，紫菜干中维生素B_<12>对大鼠具有生物利用度。杜氏藻生长对维生素B_<12>的需求量和外源维生素B_<12>的吸收量均不显著。在不含维生素B_<12>的培养基中生长的杜氏藻细胞含有生物活性维生素B_<12>（20.18±5.53 ng/g湿重）。细胞中发现的维生素B_<12>约77%是大分子组分（腺苷- B_<12>, 39%；甲基- B_<12>, 21%）。这些结果表明，该藻类具有合成维生素B_<12>辅酶的能力。研究一种红藻中非含铁氨基酸的抗氧化化合物。用Dowex柱对培养菌体的乙醇提取物紫菜（Porphyra yezoensis）进行分离。硫氰酸铁和TBARS测定表明，碱性组分V具有较强的抗氧化能力。在该碱性组分中，除鸟氨酸、赖氨酸和精氨酸外，还检测到组氨酸、3-甲基组氨酸、肌肽和鹅氨酸。肌肽和雁氨酸的出现表明组氨酸相关化合物也参与了叶斑参的抗氧化反应。采用硫氰酸铁法和硫代巴比妥酸法测定了苏沙宾提取物的抗氧化活性。氯仿-甲醇提取物的甲醇、丙酮、乙酸乙酯和依维辛烷提取物，以及氯仿-甲醇提取物的氯仿可溶性和水溶性组分的活性均高于-生育酚。热水提取物几乎没有表现出活性。薄层色谱分析表明，活性提取物中含有多种抗氧化剂。氯仿可溶性部分的活性是由于叶绿素类似物。从甲醇提取物中分离出一种强抗氧化剂，并伴有亮氨酸和苯丙氨酸等几种氨基酸。该化合物被鉴定为乌苏烯，一种类似于真菌孢素-甘氨酸的氨基酸。少

Vitamin B_<12> concentrations of dried green (Enteromorpha sp.) and purple (Porphyra sp.) Iavers (nori) were determined by both Lactobacillus leichmannii ATCC 7830 microbiological and intrinsic factor chemiluminescence methods. The values determined by using the microbiological method (68.58±2.9 and 32.26±1.61 u g/1OOg of dry weight) were identical to (bund by using the chemiluminescence method (60.20±2.21 and 25.07±0.54 <μg/100g dry weight) in both dried green and purple lavers, respectively. The vitamin B_<12> concentration of an algal health food, spirulina (Spirulina sp.) tablets, was determined. The values determined with the microbiological method were 〜6-9-fold greater in the spirulina tablets than the values determined with chemiluminescence method. The contents of vitamin B_<12> in foods were assayed by using a fully automated chemiluminescence analyzer, which was compared with a microbiological method. In shellfishes and spirulina, the values determined by the microbiological … More method were〜6-8 told greater than the values determined by the chemiluminescence method, although there was good similarity between the values by the two methods in other foods. To clarify the bioavailability of vitamin B_<12> in lyophilized purple laver(nori), total vitamin B_<12> and vitamin B_<12> analogue contents were determined, and the effects of feeding the laver to vitamin B_<12>-deficient rats were investigated. The purple laver contained five types of biologically active vitamin B_<12> compounds(cyano-, hydroxo-, sulfito-, adenosyl-, and methylcobalamin), in which vitamin B_<12> coenzymes (adenosyl- and rnethylcobalamin) comprised about 60% of the total vitamin B_<12>. When 9-week-old vitamin Burdeficient rats, which excreted substantial amounts of methylmaIonic acid(71.7(SE 20.2) μ mol/d) in urine, were fed the diet purple laver (10μg/kg diet) for 20 d. urinary methylmalonic acid excretion (as an index of vitamin B_<12> deficiency) become uncletectable and hepatic vitamin B_<12> (especially adenosylcobalamin) levels were significantly increased. These results indicate that vitamin B_<12> in dried purple laver is bioavailable to rats. Dunaliella tertiolecta neither required vitamin B_<12> for growth nor took up exogenous vitamin B_<12> significantly. The Dunaliella cells grown in a vitamin B_<12>-free medium contained biologically active vitamin B_<12> (20.18±5.53 ng/g wet weight). About 77% of vitamin B_<12> found in the cells was recovered in macromolecular fractions (adenosyl- B_<12>, 39% ; and methyl- B_<12>, 21%). These results suggest that the alga has ability to synthesize vitamin B_<12> coenxymes.To examine the antioxidative compounds of nonpmtein amino acids in a red alga. Porphyra yezoensis, an ethanol extract of the cultured thalli was fractionated with Dowex column. The basic fraction V showed strong antioxidative capacities with ferric thiocyanate and TBARS measurements. In this basic fraction, histidine, 3-methylhistidine, carnosine, and anserine wewre detected beside ornitine, lysine, and arginine by amino acid analysis. The occurrence ofcarnosine and anserine suggests that the histidine-related compounds also contribute to the antioxidative reactions in P. yezoensis. The antioxidant activities of several extracts from Susabinori were measured by the ferric thiocyanate method and the thiobarbituric acid method. The methanol, acetone, ethyl acetate, and Ivexane extracts, and the chloroform-soluble and water-soluble fractions from the chloroform-metanol extract exhibited higher activities than a -tocopherol. The hot water extract showed little activity. Thin-layer chromatography analysis of the active extracts suggested the existence of several antioxidants. The activity of the chloroform soluble fraction was due to chlorophyll analogs. A strong antioxiclant was isolated from the methanol extract, accompanied by several amino acids such as leucine and phenylalanine. This compound was identified as usujilene, a kind ofmycosporine-glycine like amino acid. Less