【作者】 管佳；

【导师】 王向阳；

【作者基本信息】 浙江工商大学 ， 农产品加工及贮藏， 2009， 硕士

【摘要】 西兰花也叫青花椰菜（Brassica oleacea）,英文名称Broccoli,是十字花科（Cruciferous）植物的一种。其硫代葡萄糖苷含量非常丰富,含有的硫苷以萝卜硫苷（也称葡萄糖莱菔子苷,4-甲基亚砜基丁基硫代葡萄糖苷,Glucoraphanin）和3-甲基亚磺酰基丙烯基硫代葡萄糖苷（Glucoiberin）为主。硫代葡萄糖苷是十字花科植物中重要的次生代谢物。它在内源芥子酶作用下水解为具有不同生理功能的活性物质。萝卜硫苷是硫代葡萄糖苷的一种,其降解产物主要以萝卜硫素（1-异硫氰酸-4-甲磺酰基丁烷,Sulforaphane,1-isothiocyanato-4-（methysulfiny）-butane）、萝卜硫腈（Sulforaphane nitrile）、嚅唑烷硫酮（Goitrin）,其中量最高的是萝卜硫素。萝卜硫素具有强抗癌活力,相对分子质量为177.29,分子式C₆H₁₁S₂NO,结构式如下:本课题以西兰花为原料,研究了西兰花中萝卜硫苷提取工艺,萝卜硫素酶解提取工艺,萝卜硫苷和萝卜硫素的免疫增强活性,萝卜硫素抗癌活性以及西兰花萝卜硫素产品的抗癌防癌作用,此外还对西兰花萝卜硫苷提取物产品的稳定性进行了研究。为了从西兰花中获得萝卜硫苷,以HPLC方法检测萝卜硫苷含量,研究了西兰花不同品种、不同部位组织、提取溶剂、原料预处理状态等条件对萝卜硫苷提取的影响。结果发现新鲜西兰花比冷冻和干燥的样品高;幼叶比花球和茎高;1:1的95%溶剂A提取4次比1:5的80%溶剂A提取2次高;提取温度70℃比常温高。为了缩短萝卜硫苷酶解时间以及了解冷冻西兰花对萝卜硫素提取的影响,得到高产量的萝卜硫素。以HPLC方法检测萝卜硫素,研究了酶解温度、酶解时间等条件对西兰花酶解产物萝卜硫素含量的影响。结果显示:新鲜西兰花:60℃水解时间6min;冷冻西兰花:温度55℃水解时间5min。本试验用纯品萝卜硫素和萝卜硫苷,对小鼠进行灌胃,以小鼠细胞免疫和体液免疫结果为指标,判断对小鼠是否起到增强免疫性功能作用,并确定其人体推荐量。结果动物实验表明,人体（50kg）每天服食萝卜硫苷34mg有增强免疫力功能作用。人体（50kg）每天服食萝卜硫素34mg有增强体液免疫作用。本试验用MTT法证明了萝卜硫素是西兰花中有效的抗肿瘤活性物质。此外本试验还用MTT法证明了自主开发的西兰花萝卜硫素提取物产品（每1g提取物含萝卜硫素1.02mg）对小鼠白血病P388细胞的体外增殖抑制活性相对较强,在200μg/mL时抑制率为51.5士2.8%。本试验研究了自主开发的西兰花萝卜硫素提取物产品（每1g提取物含萝卜硫素1.02mg）不同剂量对小鼠肿瘤的抑制作用,对免疫器官的影响及其防癌效果。方法:采用ICR小鼠,随机分组进行实验,观察西兰花萝卜硫素提取物不同剂量对小鼠肿瘤的抑瘤率的差别以及对胸腺指数、脾脏指数的影响。防癌试验采用不同剂量的西兰花萝卜硫素提取物产品进行小鼠灌胃,15天后,检测小鼠肝、肺、肾器官的谷胱甘肽S-转移酶（GSTs）活性和血液中一氧化氮（NO）含量,看其变化,以此来判断防癌的效果。实验结果:该产品动物试验显示随着灌胃剂量的上升,胸腺指数、脾脏指数明显降低。体重为18∽22克的小鼠,每天灌胃量9g/kg连续灌胃8天,对小鼠肝癌H₂₂的增长有轻微的抑制作用。此外西萝提高了小鼠肝、肺、肾的GSTs活性,降低了血液中的NO浓度,并且随西兰花萝卜硫素提取物量上升,作用效果也增加。说明西兰花萝卜硫素提取物具有有效的防癌作用。体重为30∽35克的小鼠,每天灌胃量1.85g/kg以上的西兰花萝卜硫素提取物（每1g提取物含萝卜硫素1.02mg）,连续灌胃15天,有比较好的防癌作用。更多还原

【Abstract】 Broccoli, a cruciferous plant, is also called Brassica oleacea. The glucosinolates content of broccoli is very rich .Glucoraphanin is the main composition of glucosinolates（GSs） which is a secondary metabolites are higher bioactivity compounds found in special plants. The main biological activities of glucoraphanin is Sulforaphane which is due to hydrolysis products released by the action of myrosinase.Sulforaphane can be obtained form cruciferous vegetables as one of the products of enzymatic or acid hydrolysis of glucoraphanin, which has been indentified the best anticancer activity. Molecular weight is 177..29, formula is C₆H₁₁S₂NO, structure as follows:In this paper, the optimum extraction technology getting high output of glucoraphanin from broccoli, the optimum glucoraphanin zymohydrolysis technology getting high output of sulforaphane from broccoli, the effect of sulforaphane and glucoraphanin on the immune system, effect of sulforaphane from broccoli on antitumor and preventing cancer, the stability of broccoli extract （27.9±1.2mg glucoraphanin/g） were studied, which established the groundwork for developing broccoli concentration Product.The experiment was carried out for finding out the optimum extraction technology getting high output of glucoraphanin from broccoli, the effects of solvent, varieties, parts, state, temperature, time on the extracting of glucoraphanin existed in broccoli were studied with broccoli as experiment material and the output of glucoraphanin as indicator. It was identified by HPLC. The optimized extraction processes were as follows: state: fresh broccoli; part: leave; solvent: 1:1, 95% A; extracting temperature is 70℃, extracting times is four。The experiment was carried out for finding out the optimum glucoraphanin zymohydrolysis technology getting high output of sulforaphane from broccoli, the effects of hydrolytic temperature, time on the extracting of sulforaphane [1-isothiocyanato-4-（methylsulfinyl）-butane] from glucoraphanin existed in broccoli were studied with broccoli as experiment material and the output of sulforaphane as indicator. It was identified by HPLC. The optimized extraction processes were as follows: fresh broccoli, hydrolytic temperature is 60℃, time is 6min; frozen broccoli: hydrolytic temperature is 55℃, time is 5min.The effect of sulforaphane and glucoraphanin on the immune system was studied by using BALB/c mice and the amount recommended by the human body was determining. Result: the immunomodulatory activity of glucoraphanin was indicated and the amount recommended by the human body （50kg） of glucoraphanin was 34 mg. The humoral immunomodulatory activity of sulforaphane was indicatedSulforaphane has been indentified the best anticancer activity by the experimental results of MTT. Significantly effect of broccoli extract （1.02mg sulforaphane /g） on tumor cell P338 was found with 200μg/mL.The experiment was carried out for accessing the effect of sulforaphane from broccoli on antitumor and preventing against cancer. Objective It is to observe the restraining tumor action of sulforaphane with different dosage on transplanted tumor H₂₂ of mice and the influence on immune organ. Methods ICR species mice were chosen and randomly divided into five groups. The difference of restraining tumor rate of sulforaphane with different dosage only on transplanted tumor H₂₂ of mice and the influence on thymus index, spleen index were observed. The animal test showed that the sulforaphane might have a little effect of antitumor of mice after mice （4∽6 weeks, body weight as 18∽22g） gavaged with 9g/kg dosage of broccoli extraction （1.02mg sulforaphane/g） daily for 8 days. Mice were used as animal experiments for accessing preventive cancer effect of sulforaphane. The mice were gavaged with the different dosage of broccoli extraction （maily active composition is sulforaphane） daily for 15 days. Then the nitric oxide （NO） content in serum and the glutathione S-transferase （GSTs） activities in liver, lung and kidney of mice were measured for accessing the effect of preventive cancer. The animal test showed that the NO content in serum of mice was reduced and GSTs activities was enhanced in liver, lung and kidney of mice with gavage dosage of sulforaphane increase. This indicated that the sulforaphane might have high effect of protecting against cancer of mice. Significantly effect of preventing against cancer was found after mice （12∽16 weeks, body weight as 30∽35g） gavaged with 1.85g/kg dosage of broccoli extraction （1.02mg sulforaphane /g） daily for 15 days.更多还原