忍冬科接骨木属(Sambucus L.)植物大部分分布在地球北半球的温带和亚热带地区,其品种可达20余种。我国的接骨木广泛种植在我国的东北、西北至甘肃、西南至云南等地区,本土品种有4种~5种,从国外引种栽培了1种~2种,如毛接骨木、血满草和接骨草等[1-3]。
接骨木,又名接骨丹、大接骨丹、铁骨散等,在5月~6月开花,9月~10月结果[4-5]。该植物含有糖及糖苷类、黄酮类、萜类、酚及酚苷类等生物活性物质,在世界范围内作为功能性成分广泛用于食品、制药、绿化以及洗护用品等[6-8]。接骨木根、茎、叶、花、果实均可入药,用于治疗类风湿关节炎、痛风、骨折等疾病[9],具有预防骨质疏松、抗病毒、抗炎、抗氧化、抗肿瘤、抗艾滋病、降血糖、降血脂等[9-13]功能;另外接骨木还被用于制作各种饮料、果酱、果冻、果酒、调味品、眼霜以及洗发水等,同时还具有较高的观赏价值[6,14-18]。本文对接骨木的活性成分及其功能进行综述,旨在为接骨木的合理开发和综合利用提供理论基础。
多糖(又叫多聚糖)是一类广泛分布于自然界且具有多种生物活性的天然高分子化合物,是由10多个单糖(醛糖或酮糖)以α-糖苷键或β-糖苷键连接而成的碳水化合物聚合物,是生命有机体的重要组成部分,对维持机体正常生命活动至关重要,分子量可达数万以上[19-20]。从植物、动物以及微生物中提取的多糖具有抗氧化、抗肿瘤、抗疲劳、抗炎及抗菌等生物活性,又能控制细胞的分裂、分化,调节细胞的生长与衰老[21-25]。
研究表明,从接骨木中提取、纯化、冷冻干燥后的多糖为亲水性淡黄色粉末。吴志诚[26]采用传统热水提取法提取接骨木茎中的粗多糖,以温度、料水比、时间为单因素设计试验,得出料水比 1∶50(g/mL),在100℃温度下浸提4 h为接骨木茎粗多糖的最佳提取工艺;分析了分别采用水提法、酸提法、超声波辅助提取法、超声波催化纤维素酶提取法提取接骨木叶粗多糖,提取率分别为7%、5.32%、6.78%、2.89%;并对提取的粗多糖进行纯化,按 1∶40(g/mL)在 100℃水中提取 3 h,加酶辅助,木瓜蛋白酶-TCA法(多糖溶液∶正丁醇=1∶1,体积比)脱蛋白质,多糖溶液与无水乙醇体积比为1∶4醇沉,接骨木粗多糖得率为15.36%。刘莹等[27]研究表明,以1∶20(g/mL)的料液比,在85℃条件下提取3 h,接骨木茎粗多糖的提取率可达10.21%;之后刘莹等[28]以接骨木叶为原料,得到75℃条件下,以1∶30(g/mL)料液比,提取3 h为接骨木叶粗多糖提取的最佳工艺,提取率可达15.52%;选择石油醚、过氧化氢、木瓜蛋白酶法-三氯乙酸正丁醇联合法对接骨木叶粗多糖进行纯化,纯化后蛋白质、糖、水分、灰分在接骨木叶粗多糖中的含量分别为5.38%、33.62%、12.56%、1.70%[29]。
黄酮类化合物是一类具有多种生理功能的天然生物活性成分,其特征是由一个杂环六元吡喃酮环连接的两个苯环,广泛存在于水果、蔬菜和其他粮食作物中,是人类日常饮食中的重要组成部分,具有抗氧化、抗菌、抗肿瘤、抗病毒、降血脂等生物活性[30-34]。
苏新芳等[13]用超声波辅助提取接骨木叶总黄酮,结果表明最佳工艺参数为超声频率40 kHz、超声功率500 W、料液比 1∶40(g/mL)、提取温度 70℃、提取时间1 h,此时总黄酮提取率为18.67%;柳佳莹[35]在单因素试验的基础上设计响应面试验,得出接骨木叶总黄酮的最佳提取工艺:温度70℃、时间1 h、乙醇体积分数70%、料液比 1∶70(g/mL),此条件下得率为 7.25%,纯度为31.3%,后经D101型大孔树脂纯化,纯度可达70.30%;张爽等[36]采用响应面法优化接骨木叶总黄酮的提取工艺,得出接骨木叶总黄酮的提取量在料液比1∶19.53(g/mL),乙醇体积分数 76.03%,超声温度40℃,超声时间48.86 min条件下最大,为5.80 mg/g,超声温度对接骨木总黄酮提取率影响最小;李安林等[37]采用乙醇浸提法,得出提取温度对接骨木茎总黄酮提取率的影响最大,乙醇浓度影响最小,以料液比1∶20(g/mL),乙醇浓度 95%,提取温度 80℃,提取时间1 h为接骨木茎总黄酮的最佳提取工艺参数提取总黄酮,提取率可达1.37%。
花青素是一类带有氧鎓离子的特殊水溶性天然植物色素,多以苷的形式广泛存在于忍冬科植物的果实、根、茎、叶中[38-39]。其稳定性较差,易受氧、温度、光、酶、pH值影响,在高温条件下易降解,而在酸性条件下稳定[40-41],具有抗氧化、抗衰老、抗癌、抗炎、抗肿瘤、有效清除自由基和防止心血管疾病等[42-45]功能,特别是在抗氧化和清除自由基能力方面相当于维生素C的20倍,因其安全无毒,被称为目前最安全、最高效的天然抗氧化剂[46-47]。
于洪超等[48]以接骨木果实为原料,选用超声波辅助提取法,得出料液比1∶40(g/mL),乙醇体积分数69%,提取温度49℃,提取时间0.5 h为接骨木花青素的最佳提取工艺,得率为2.54%。冯文娟等[49]以接骨木鲜果、干果为原料,采用乙醇浸提法提取花青素,此条件下花青素得率分别为0.43%和1.52%,后经分离技术、正己烷萃取、大孔吸附树脂(AB-8型)纯化后,花青素得率分别为27.74%、29.32%,且正己烷纯化花青素的效果较好。
接骨木具有多种生物活性,如Veberic等[50]从接骨木果实中鉴定出4种有机酸:柠檬酸、苹果酸、莽草酸和延胡索酸;在槲皮素组中鉴别出槲皮素、槲皮素3-芦丁苷和槲皮素3-葡萄糖苷,采用高效液相色谱发现有5个花青素峰。杨文松[51]以正丁醇萃取的接骨木花为研究对象,采用硅胶、聚酰胺柱、Sephadex LH-20等层析手段分离纯化,在薄层色谱、核磁共振、质谱、红/紫外光谱等条件下,分离得到14个化合物及13个化合物结构。另外,迄今为止,从接骨木属植物分离出11个酚及酚苷类化合物,56个苯丙素类化合物,51个萜类化合物,35个黄酮类化合物以及13个生物碱类化合物等[2]。
接骨木具有较高的抗氧化活性,主要与接骨木果实的化学结构和组成有关,其富含酚类物质,对DPPH自由基的清除率约为82%~89%。李安林等[37]采用乙醇浸提法提取接骨木茎总黄酮,得出接骨木茎总黄酮对DPPH自由基活性的半数抑浓度IC50为23.81 μg/mL,表明总黄酮具有较强的抗氧化活性;冯文娟等[49]采用同样的方法提取接骨木果实花青素,得出其质量浓度与花青素对DPPH自由基清除能力成正比关系,在花青素质量浓度为200 μg/mL时,DPPH自由基的清除率为89.40%,相当于维生素C的96.28%;柳佳莹[35]将从接骨木叶中提取的总黄酮按照低(80.5 mg/kg)、中(161 mg/kg)、高(322 mg/kg)3 个剂量组,灌入小鼠胃中,再对小鼠的脏器、血液、股骨进行药理研究,得出接骨木叶总黄酮具有良好的抗氧化、抗骨质疏松作用。Xiao等[14]从接骨木中分离出的香草酸,能刺激UMR106细胞增殖,增强碱性磷酸酶活性,改变成骨细胞功能和破骨细胞形成相关mRNA基因的表达,证实了接骨木具有抗氧化、预防骨质疏松症、刺激碱性磷酸酶的活性、调节成骨细胞和破骨细胞等生理作用;Xiao等[52]以9个月的雌性鼠为模型,将去卵巢或假性手术的老龄鼠分组,口服给予低、中、高剂量接骨木12周,研究结果表明,接骨木中的木质素能有效维持大鼠骨小梁和骨皮质的健康、调节成骨细胞和破骨细胞的形成过程,具有较强的抗骨质疏松活性,同时在大鼠体内具有抗氧化作用,在体外具有抑制血清合成的作用。
Roschek等[53]研究表明,从接骨木果实中提取的类黄酮是抗流感活性的重要组成部分,这些类黄酮分子通过合成5,7,30,40-4-O-甲基槲皮素和外消旋杨梅素与H1N1病毒粒子结合,在体外阻断H1N1感染,阻止H1N1病毒进入宿主细胞,结合后的化学物质再与H1N1病毒结合,阻止宿主细胞进入或识别,进而在体外抑制H1N1感染,达到抗病毒作用。雷用东等[11]以接骨木为原料,用提取物花色苷感染猪传染性胃肠炎病毒(transmissible gastroenteritis virus,TGEV) 的 猪 睾 丸(swine testicular,ST)贴壁细胞,研究表明花色苷浓度与ST存活数成正比关系,当接骨木花色苷浓度为50 μg/mL时,TGEV的活性最大,表明从接骨木中提取出的花色苷能有效抑制TGEV繁殖;而Torabian等[42]研究表明,高浓度的花青素能完全抑制流感病毒活性,低浓度花青素抑制抗流感病毒活性的能力较弱,其作用机理有:直接抑制病毒进入;抑制病毒从细胞到细胞的传播;调节细胞因子的释放,间接抑制病毒进入。另外,Neves等[54]从接骨木中提取花青素,通过无细胞试验证实,花青素对鱼藤酮诱导的毒性神经具有保护作用,通过增强酶的活性及细胞的抗氧化防御能力对鱼藤酮等氧化毒性刺激提供神经保护作用。Weng等[55]指出接骨木茎乙醇提取物和一些酚酸(咖啡酸、绿原酸和没食子酸)成分具有抗HCoV-NL63病毒活性。文献[56-57]研究表明从接骨木中分别提取得到的落叶松树脂醇、(+)-皮树脂醇都具有抗真菌作用。赵淑红等[58]从接骨木中提取的齐墩果酸、熊果酸也同样具有抗病毒活性,可以有效治疗急性病毒性肝炎。
王丹蕊等[59]研究多糖对大鼠糖尿病的调节作用,对几种不同来源的植物多糖进行比较,对雄性SD大鼠腹腔注射链脲菌素,设置对照试验,利用动态血糖测量方法得出结论,接骨木多糖能有效降低链脲佐菌素诱发糖尿病大鼠的血糖,同时血浆内高密度脂蛋白水平明显升高。Lv等[60]采用高压液法提取接骨木种子油,用尿素包合法分离出亚油酸,采用不同浓度的亚油酸饲喂高脂血症小鼠,通过亚油酸对α-葡萄糖苷酶抑制作用的测定,得出接骨木中的亚油酸具有良好的降血脂、降血糖活性。
研究表明,接骨木除了具有抗氧化、抗骨质疏松、抗炎、抗病毒、抗真菌、降血糖、降血脂活性外,还具有抗肿瘤、抗衰老、促进骨折愈合、增强免疫以及影响成骨细胞增殖和分化等活性[9-13]。努尔曼·衣西坚等[61]采用酶联免疫法,证明了接骨木提取物能增强大鼠骨代谢情况,能有效促进大鼠骨细胞和破骨细胞的合成,增强成骨细胞和破骨细胞活性,促进骨折愈合。宋琳亮等[62]采用热水浸提法提取的接骨木茎多糖喂养大鼠,确定多糖对大鼠胰岛细胞活力、细胞活性(四氧嘧啶感染后)以及接骨木茎多糖和四氧嘧啶共同对大鼠胰岛细胞分泌胰岛素功能的影响,结果表明接骨木茎多糖能促进大鼠胰岛细胞的增殖,避免大鼠胰岛细胞遭受四氧嘧啶损伤。
接骨木作为一种植物,分布广泛,营养价值极高,含有多种生物活性成分,除了具有预防骨质疏松、抗病毒、抗炎、抗氧化、抗肿瘤等作用,还具有护肝、保护神经、抗抑郁等多种生理功能,并且在养生、饮食以及绿化美观等方面被广泛使用。近几年来,国内学者对接骨木抗炎、促进骨折愈合等功能、接骨木的根皮和茎皮的化学成分、药理作用以及对接骨木的生物活性物质进行提取分离、纯化,发现接骨木在生理药理功能方面的开发潜力巨大。但目前接骨木生物活性物质以基础研究为主,对其结构鉴定及药效机理的研究尚缺乏深入和系统的研究。
因此,加深对接骨木生物活性物质的高效提取、分离纯化、结构鉴定和药理机理的研究,建立多种简单有效的实验模型,对接骨木各部分在临床上的应用,开发深层次接骨木药品,促进接骨木深加工、产业发展以及提高农产品附加值、提高农民收入、预防疾病、促进人类健康等具有十分重要的意义。
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