长白山野生膜荚黄芪不定根总黄酮积累规律研究(Ⅰ)

以膜荚黄芪不定根为试材,测定了其不同生长时期不定根的生物量、总黄酮含量和体外抗氧化活性。结果表明:不定根生物量呈"S"型曲线变化,总黄酮的积累有2个波峰,抗氧化活性有1个波峰;生长至40d时,生物量、总黄酮含量和自由基清除率最高,分别为0.513g、2.048mg/g和92.29%,均显著高于其它时期;因此,确定40d为收获总黄酮的最佳时期。     

氮和磷对膜荚黄芪不定根芒柄花苷积累的影响

以膜荚黄芪不定根为试材,通过改变B5培养基成分,依次研究了总氮含量、NH⁺₄∶NO^-₃比例、磷酸盐含量对不定根生物量以及芒柄花苷积累的影响,以期得到最佳氮、磷配比。结果表明:当培养基中总氮浓度为3.0 mmol·L^-1、NH⁺₄∶NO^-₃比例为1.5∶1.5、PO₄^3-浓度为0.625 mmol·L^-1时,黄芪不定根生物量及芒柄花苷含量均依次达到了最大值,且与初始培养基相比芒柄花苷的含量分别增加了7、14、19倍。因此,该方法简单可靠,且配比结果为进一步利用膜荚黄芪不定根规模化生产芒柄花苷奠定了理论基础。     

膜荚黄芪子叶节植株再生体系的建立

以膜荚黄芪无菌苗子叶节为外植体,建立了简单、高效和稳定的膜荚黄芪再生体系。结果表明:不定芽诱导的最适培养基是MS+6-BA 1.0mg/L+NAA 0.10mg/L,每个子叶节外植体平均出芽数为3个以上,再生频率达75.5%;生根最适培养基为1/2MS+IBA 0.5mg/L;移栽后成活率达75%以上。     

膜荚黄芪种子中花青素提取方法的优化

以膜荚黄芪种子为试材,在单因素试验基础上,采用L25(56)正交实验法,以吸光度为评价指标,优化了花青素提取方法,为膜荚黄芪的开发利用提供参考依据。结果表明:料液比是对花青素提取效果影响最大的因素,最佳提取方法为料液比1∶8g·mL-1、甲醇体积分数85%、加酸量1.0%、超声时间30min、超声温度30℃,该提取方法简单可靠,具有较高的抗氧化能力。     

长白山区野生膜荚黄芪种质资源的SRAP分析

以野生膜荚黄芪的叶片为试材,采用SRAP分子标记技术,研究了长白山区野生膜荚黄芪种质资源的遗传多样性,以期为膜荚黄芪种质资源的保存和优良品种的选育奠定基础。结果表明:从300对引物组合中选出扩增产物条带多,特征性强,重复性明显的16对引物组合;利用这些引物进行PCR扩增,共得到129条条带,其中多态性条带数为95条。聚类分析结果表明,长白山区野生膜荚黄芪种质资源之间存在着丰富遗传差异。     

膜荚黄芪查尔酮还原酶基因的克隆与表达分析

从膜荚黄芪中克隆了CHR基因,GenBank登陆号为KY086286(AmCHR),AmCHR全长cDNA为1 173 bp,开放阅读框为957 bp,编码318个氨基酸,其分子量为35.58 kDa,等电点为6.26。生物信息学分析表明AmCHR蛋白与豆科植物CHR同源,无信号肽,定位于细胞质。实时荧光定量PCR和高效液相色谱分析结果显示AmCHR在根、茎、叶中的表达水平与毛蕊异黄酮葡萄糖苷的含量变化一致。该研究从膜荚黄芪中克隆了1个新的AmCHR基因,推测AmCHR的表达与膜荚黄芪毛蕊异黄酮葡萄糖苷的积累密切相关。     

长白山野生膜荚黄芪cDNA文库的构建与初步鉴定

以长白山区野生膜荚黄芪根和叶为试材,参照TRIzoL说明书提取总RNA,采用SMART技术构建其全长cDNA文库,为分子水平上揭示膜荚黄芪次生代谢调控机制提供充足的基因资源。结果表明:原始文库滴度为4.8×105 cfu/mL,重组率接近99.1%,插入片段大于0.5kb,平均大小1.54kb。所构建的cDNA文库的库容量、克隆效率和全长率满足挖掘膜荚黄芪次生代谢相关基因的要求。     

不同基质对膜荚黄芪根系生长发育的影响

以膜荚黄芪为试材,采用人工栽培的方法,在不同体积配比的各混合基质中培养,测定其株高、根长和根鲜重,研究不同基质对黄芪根系生长发育的影响。结果表明:不同基质对黄芪株高影响不显著,对根长和鲜重的影响作用显著。Ⅳ号基质为黄芪的首选混合基质。     

青天葵总黄酮抗氧化及抑菌活性

以青天葵为试材,乙醇为溶剂,对青天葵进行总黄酮提取;采用清除羟自由基、DPPH自由基、超氧阴离子自由基和测定总还原能力的方法来评价青天葵总黄酮抗氧化活性;并通过测定其对6种试验菌的最低抑菌浓度来考察其抑菌活性。结果表明:青天葵黄酮提取液对羟自由基的清除优于同浓度的抗坏血酸溶液,清除DPPH自由基的IC50是17.4μg·mL~(-1),清除超氧阴离子自由基的IC50是95.5μg·mL~(-1),还原能力稍弱于抗坏血酸;对白色念珠球菌、铜绿假单胞菌和肺炎克雷伯氏菌的抑制作用较强,最低抑菌浓度均为0.25mg·mL~(-1)。     

低温对黄瓜幼苗膜脂过氧化的影响

研究了低温对黄瓜幼苗膜脂过氧化的影响。结果表明：低温下细胞的膜脂过氧化程度明显加剧,丙二醛（ＭＤＡ）含量显著增加;过氧化物酶（ＰＯＤ）活性上升,过氧化氢酶（ＣＡＴ）活性明显下降;ＭＤＡ含量与黄瓜幼苗耐低温能力呈极显著负相关,ＰＯＤ和ＣＡＴ活性与黄瓜幼苗耐寒性分别呈正相关和显著正相关。     

微波提取黄芪黄酮的方法研究

以黄芪为试材,用微波消解仪提取黄酮,分光光度法测定黄酮提取率,采用单因子试验及正交实验研究黄芪黄酮的最佳提取条件。结果表明:乙醇浓度95%,提取时间20min,料液比1∶15g/mL,提取温度90℃时,提取效果最好,黄酮提取率为0.489%,各因子对提取效果影响从大到小依次为提取温度乙醇浓度料液比提取时间。     

紫萼玉簪根中总黄酮提取工艺优化

以紫萼玉簪根为试验材料,采用单因素结合正交实验方法,研究紫萼玉簪根中总黄酮的最佳提取工艺对玉簪属植物的影响。结果表明:超声波提取法提取紫萼玉簪根中总黄酮的最佳提取工艺为:乙醇浓度80%,提取温度80℃,提取时间60min,料液比1∶15,总黄酮得率25.48mg/g。     

黄芩总黄酮含量的累积规律研究

以一年生和二年生黄芩为试材,研究了不同部位、不同生长时期总黄酮含量的累积规律,以期为黄芩的种植和确定最佳采收时期提供参考。结果表明:黄芩总黄酮含量"根叶茎";在一年生和二年生黄芩中的累积规律不同,其中一年生黄芩根总黄酮含量在9月份最高、达到46.97mg/g,二年生黄芩根总黄酮含量在6月份最高、达到54.50mg/g,显著高于其它时期;因此,确定一年生黄芩最佳采收时期为9月份,二年生黄芩最佳采收时期为6月份。     

淹水胁迫对黄瓜幼苗生长及不定根解剖结构的影响

以黄瓜幼苗为试材,研究了淹水胁迫(0、4、8、12d)对黄瓜幼苗生长指标及不定根解剖结构的影响。结果表明:淹水胁迫抑制了黄瓜幼苗的根、茎、叶鲜质量、株高及子叶上下1cm处茎粗的正常生长,淹水胁迫4d对根、茎、叶鲜质量及株高产生显著抑制作用,淹水8、12d分别显著抑制了子叶上、下1cm处茎粗生长,而淹水胁迫12d内未对黄瓜幼苗叶片数的生长起到抑制作用。黄瓜幼苗叶片MDA含量在淹水胁迫4d后淹水处理显著高于对照。淹水胁迫12d后,黄瓜不定根皮层出现溶生性通气组织,但不发达。说明黄瓜幼苗具有一定的耐淹水能力,但能力有限。     

外源水杨酸对低温胁迫下黄瓜幼苗根系脂肪酸不饱和度的影响

为了阐明水杨酸(SA)诱导黄瓜幼苗耐冷性的作用机理,以28℃/18℃(昼/夜)为对照,测定了外源SA对低温胁迫(8℃/8℃)下根系脂肪酸总量、组分、不饱和度以及脂肪酸去饱和酶基因(CsFAD)表达水平的影响。结果表明:低温提高了根系总脂肪酸含量和饱和脂肪酸组分C16︰0、C18︰0的相对含量,并降低了多不饱和脂肪酸C18︰3的比例,根系脂肪酸不饱和度和双键指数降低;而低温下外源施用SA后,脂肪酸组分的变化得到显著缓解,饱和脂肪酸C16︰0的相对含量降低,不饱和脂肪酸C18︰3相对含量升高,脂肪酸不饱和度升高。进一步分析CsFAD的表达水平,发现低温胁迫下,根系CsFAB2.1、CsFAB2.2和CsFAD5表达受到抑制,CsFAD3和CsFAD7的表达升高;施用SA可以解除低温对CsFAB2.1、CsFAB2.2、CsFAB2.3的抑制,CsFAD2.1和CsFAD3的表达也有所升高。低温胁迫下外源施用SA可以诱导黄瓜幼苗根系FAD基因表达,提高脂肪酸不饱和度,提高幼苗耐冷性。     

The Development of Adventitious Roots on Hardwood Cuttings of Gooseberries

Changes in percentage dry weight and in carbohydrate levels and distribution were followed in Golden Brown Lockyer onion bulbs for three months at storage temperatures of 4°, 15°, 25° and 37°C. The percentage dry weight within the bulbs increased from the outer to the inner leaf bases. Storage temperature and length of storage did not influence water loss. Sucrose concentrations increased from the outer leaf bases to become highest in the inner leaf bases, and in all leaf bases were higher at higher temperatures. Glucose and fructose levels also tended to increase from outer to inner leaf bases. After eight weeks fructose levels increased rapidly at the lower storage temperatures and this is attributed to low-temperature hydrolysis of fructans. The only fructan detected was the trisaccharide fraction, which was barely detectable in outer leaf bases and maximal in the inner ones. Trisaccharide levels were generally lower at the lower temperatures, thus supporting the suggestion of low-temperature hydrolysis of fructans. No information was obtained regarding storage temperature effects on fructans having a higher degree of polymerization than the trisaccharide. The results are discussed relative to temperature and time and the development of storage rots.     

Astragalus membranaceus inhibits bleomycin-induced pulmonary fibrosis in mice by antioxidation

AIM:To investigate the effect of Astragalus membranaceus on the balance of oxidation and antioxidation in bleomycin-induced pulmonary fibrosis in mice and the possible anti-fibrosis mechanism of Astragalus membranaceus.METHODS:Female KM mice (n=36) were randomly divided into 3 groups.The mice in control group were administered with saline aerosol intratracheally.The mice in fibrosis group were administered with bleomycin at dose of 3 mg/kg aerosol intratracheally.The mice in Astragalus membranaceus group were administered with bleomycin at dose of 3 mg/kg aerosol intratracheally and then intraperitoneal injected with Astragalus membranaceus parenteral solution at daily dose of 1.7 g/kg.All mice were sacrificed 14 d after the treatment,and the lungs and serum were collected for detection.Hematoxylin-eosin staining was performed in the lung tissue.The mRNA expression of superoxide dismutase 1/2/3(SOD1/2/3),catalase (CAT),NADPH oxidase 2/4(NOX2/4) and α-smooth muscle actin (α-SMA) was detected by RT-PCR,and the protein expression of α-SMA and NOX2/4 was determined by Western blot.The concentration of malondialdehyde (MDA) and total antioxidant capacity (T-AOC) in the serum were measured by a colorimetric method.RESULTS:The pathological injury was obviously observed in bleomycin group compared with control group,but was attenuated in Astragalus membranaceus group.The α-SMA mRNA and protein expression,MDA/T-AOC,NOX2,NOX4 and SOD3 mRNA expression,and NOX2 protein expression in bleomycin group were significantly higher than those in control group,while those in Astragalus membranaceus group were significantly lower than those in bleomycin group.The protein expression of NOX4 in bleomycin group was significantly lower than that in control group,while that in Astragalus membranaceus group was higher than that in bleomycin group.The mRNA expression of SOD1 and CAT in Astragalus membranaceus group and bleomycin group were decreased compared with control group.No significant difference of SOD2 mRNA expression among the 3 groups was observed.CONCLUSION:Astragalus membranaceus inhibits bleomycin-induced pulmonary fibrosis in mice by maintaining the balance of oxidation and antioxidation.