羟基自由基引起的脱氧核糖核酸损伤研究

以鲱鱼精脱氧核糖核酸(Herring sperm DNA)为研究对象,利用紫外光(UV,200～275 nm,66.4 Lx)激发纳米TiO2发生光催化作用介导产生羟基自由基(Hydroxyl radical,.OH),探讨.OH引发DNA氧化损伤特性。采用凝胶电泳和高效液相色谱(HPLC)分析法跟踪DNA损伤历程;应用电子自旋共振(Electron spinresonance,ESR)及分光光度法跟踪损伤过程氧化物种及H2O2相对浓度的变化;运用生物标准样8-羟基脱氧鸟苷(8-Hydroxy-2’-deoxyguanosine,8-OHdG)为内标物,通过HPLC分析DNA损伤产物,研究DNA损伤机理。结果表明,较单纯UV辐照或暗光(Dark)催化条件,DNA浓度10 mg/L,TiO2浓度1.5 g/L、pH 7～8,紫外光激发纳米TiO2介导产生.OH引发DNA损伤程度最大;DNA损伤为.OH氧化历程,并伴随有深度氧化过程;DNA结构中鸟嘌呤最易氧化损伤,8-OHdG为DNA氧化损伤中间产物及鸟嘌呤氧化损伤的特异产物。     

柚皮中总黄酮的提取及对羟自由基的清除作用

为充分利用柚皮资源,避免资源的浪费,探讨柚皮中总黄酮的提取、鉴别及对羟自由基清除作用,采用超声波乙醇浸提法从柚皮中提取黄酮类物质,并对所提取的黄酮类物质进行了验证和用分光光度法测定了其含量,对柚皮中总黄酮对羟自由基的清除作用进行了试验。结果表明,测得样品中总黄酮的含量为0.340 0 mg/mL,回收率为99.7%,其纯度和产率均较高。该法采用全物理过程,无任何污染,是提取柚皮中黄酮类物质的有效途径。柚皮中总黄酮提取液对Fen-ton体系产生的.OH自由基有很好的清除作用。     

红花黄色素抗氧化活性研究

通过考察红花黄色素(SY)及其主要化学成分对羟基自由基介导2-脱氧核糖氧化降解的抑制作用,以及对1,1-二苯基-2-苦肼基自由基(DPPH·)的清除能力,探究SY的体外抗氧化活性及其抗氧化的主要有效成分.通过Fenton反应产生羟基自由基,用紫外分光光度计检测了SY及其主要化学成分对2-脱氧核糖降解的抑制作用和对DPPH·的清除能力,同时对红花黄色素B(SYB)抑制2-脱氧核糖降解的作用机制做了初步研究.结果表明SY抑制Fenton反应对2-脱氧核糖的氧化降解的IC50为256.79 μg/mL,对DPPH·清除的IC50值为27.15μg/mL.羟基红花黄色素A(HSYA)和SYB是SY中抗氧化的两个有效成分,抑制2-脱氧核糖的氧化降解的IC50分别为220.68 μg/mL和207.01μg/mL;对DPPH·清除的IC50值分别为55.81 μg/mL和41.25μg/mL.SYB对2-脱氧核糖氧化降解的抑制作用机理研究表明,SYB除了对Fenton反应产生的羟基自由基具有直接清除作用外,又可通过与Fe2+离子的络合作用而阻断Fenton反应产生羟基自由基.由此可知SY具有明显的体外抗氧化活性,SYB和HSYA为其主要抗氧化活性成分.     

紫外可见分光光度法测定田七花中总黄酮含量

建立了分光光度法测定田七花中黄酮含量的方法.采用分光光度法,以芦丁为对照品,在波长510nm处对样品中的总黄酮进行含量测定.样品总黄酮在浓度0～50μg/mL范围内线性关系良好,其回归方程为A=11.069C-7.143×10-4;R=0.9993.该方法可作为田七花中总黄酮含量的测定的方法之一.     

蛇床子不同炮制品中的总黄酮含量测定

采用超声波提取法提取蛇床子不同炮制品中总黄酮,以芦丁为对照品,紫外分光光度法测定总黄酮的含量,标准曲线在2.1～11.0μg/mL范围内,吸光度与浓度的线性关系良好(R~2=0.999 7),实验的精密性和稳定性RSD分别为1.13%和2.15%,平均回收率为105.81%,RSD为2.98%。结果表明紫外分光光度法测定蛇床子不同炮制品中的总黄酮含量操作简便、结果准确、重现性好,实验方法切实可行。     

牛大力总黄酮提取工艺及不同萃取物的抗氧化活性研究

为了考察牛大力乙醇提取物中总黄酮的含量及抗氧化活性。通过L16(45)正交实验,超声波辅助提取牛大力中总黄酮,得到最佳工艺,再测试乙醇提取物和四个萃取物对羟基自由基(.OH)和DPPH自由基(DP-PH.)的清除效果。最佳工艺为:φ(EtOH)=75%、m(牛大力,g)∶V(EtOH,mL)=1∶25、温度60℃、时间60min,该条件下,牛大力总黄酮得率可达2.14mg.g-1。其中,牛大力乙醇提取物中氯仿萃取物中黄酮含量最高,为5.52mg.g-1;而且氯仿萃取物对DPPH.的清除效果最好,其半数抑制浓度(IC50)为40.97μg.mL-1;乙酸乙酯萃取物对羟基自由基的清除效果最好,其IC50值为90.5μg.mL-1。牛大力乙醇提取物中石油醚、氯仿、乙酸乙酯萃取物都有很好的抗氧化活性,且稳定性、重复性好。     

火棘果总黄酮的提取和纯化研究

采用溶剂提取法、索氏提取法、循环超声提取法提取火棘果总黄酮,利用静态吸附方法筛选分离火棘果总黄酮的最适大孔树脂,利用动态吸附方法研究最适大孔树脂纯化火棘果总黄酮的条件,并用紫外分光光度法测定其含量。得出结果为循环超声提取时间短、效率高;D101大孔吸附树脂纯化效果最好,最佳工艺为上样浓度为0.899 8g·L-1,上样液pH为4,上样体积5BV,上样速率2.5mL·min-1,用5BV70%乙醇以2.0 mL·min-1速率洗脱,经树脂纯化后总黄酮的纯度由原来的9.00%提高至28.11%。     

氮氧自由基对白血病细胞DNA合成的抑制

4-异硫氰酸盐-2,2,6,6-四甲基哌啶-1-氧自由基对白血病细胞7712的生长及DNA合成都有强烈抑制作用,对DNA合成的半抑制浓度为2.2μg/ml,在浓度为50μg/ml时抑制率高达99.7％,对小白鼠腹腔注射的半致死剂量为277mg/kg,不损伤DNA复制模板。它对DNA合成的抑制作用强于异硫氰酸盐及氮氧自由基单独作用之和。其它氮氧自由基类化合物也有较弱的抑制作用。但氮氧自由基的还原物羟胺却没有抑制作用。对致癌和抗癌的自由基机理进行了讨论。     

紫外分光光度法测定蒙药漏芦花中总黄酮含量

用紫外分光光度法测定蒙药漏芦花中总黄酮含量.以芦丁为对照品,在波长510nm处对样品中的总黄酮含量进行测定.样品总黄酮在浓度0～50μg/mL范围内线性关系良好,其回归方程为:A=0.0128C+0.0064,R=0.9995;平均加样回收率98.54%,RSD为1.320%(n=5).该方法稳定、简便、准确,可用于漏芦花中总黄酮含量的测定.     

植物多糖清除羟自由基作用的研究

研究当归、黄芪、鬼臼和猫人参的多糖对羟自由基的清除作用.采用分光光度法测定羟自由基清除率,并计算出半数清除率(IC50).当归多糖、黄芪多糖、鬼臼多糖对羟自由基的IC50分别为1.38 mg/mL、0.485 mg/mL、0.1 mg/mL,猫人参的最大清除率仅为36.84%.其清除能力鬼臼多糖＞黄芪多糖＞当归多糖＞猫人参多糖.鬼臼多糖、黄芪多糖均具有较强的清除自由基作用,且清除能力与浓度呈明显的正相关.     

The Optimization of Extraction Process,Antioxidant, Whitening and Antibacterial Effects of Fengdan Peony Flavonoids

Flavonoids have important biological activities, such as anti-inflammatory, antibacterial, antioxidant and whitening, which is a potential functional food raw material. However, the biological activity of Fengdan peony flavonoid is not particularly clear. Therefore, in this study, the peony flavonoid was extracted from Fengdan peony seed meal, and the antioxidant, antibacterial and whitening activities of the peony flavonoid were explored. The optimal extraction conditions were methanol concentration of 90%, solid-to-liquid ratio of 1:35 g:mL, temperature of 55 °C and time of 80 min; under these conditions, the yield of Fengdan peony flavonoid could reach 1.205 ± 0.019% (the ratio of the dry mass of rutin to the dry mass of peony seed meal). The clearance of Fengdan peony total flavonoids to 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical, hydroxyl radical and 2,2’-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) free radical could reach 75%, 70% and 97%, respectively. Fengdan peony flavonoid could inhibit the growth of the Gram-positive bacteria. The minimal inhibitory concentrations (MICs) of Fengdan peony flavonoid on S. aureus, B. anthracis, B. subtilis and C. perfringens were 0.0293 mg/mL, 0.1172 mg/mL, 0.2344 mg/mL and 7.500 mg/mL, respectively. The inhibition rate of Fengdan peony flavonoid on tyrosinase was 8.53–81.08%. This study intensely illustrated that the antioxidant, whitening and antibacterial activity of Fengdan peony total flavonoids were significant. Fengdan peony total flavonoids have a great possibility of being used as functional food materials.     

Response surface optimized ultrasonic-assisted extraction of flavonoids from Sparganii rhizoma and evaluation of their in vitro antioxidant activities

An efficient ultrasound-assisted extraction technique was employed to extract total flavonoids from Sparganii rhizoma. The optimum extraction conditions for the highest yield of total flavonoids were ethanol concentration 53.62%, ultrasonication time 29.41 min and ultrasound power 300 W, which were determined using response surface methodology. The extraction yields of the optimal ultrasound-assisted extraction were higher than using conventional extraction. The crude extract was then purified on a polyamide resin, whereby the flavonoids content in the purified extract increased to 94.62%. The antioxidant activities of the purified flavonoids including DPPH radical scavenging activity, ABTS+ radical scavenging activity, reducing power, hydroxyl radical scavenging activity and superoxide anion scavenging activity, were evaluated in vitro, which suggested that the flavonoids showed significant antioxidant activities. Rutin, kaempferol and formononetin were identified in the extract by comparing relative retention times and UV-Vis spectra with those of reference standards.     

明日叶黄酮类化合物清除羟基自由基活性研究

为了研究明日叶黄酮类化合物对羟基自由基的清除作用,以明日叶（主要取叶片）为原料,用体积分数为65％乙醇提取明日叶总黄酮,测定其总黄酮含量.通过Fenton反应体系产生羟基自由基,利用明日叶提取液中的功能成分黄酮类化合物对羟基自由基的清除作用进行研究.结果表明：明日叶提取物总黄酮质量分数为10.18％,且黄酮类化合物对羟基自由基有较强清除效力,当提取物总黄酮浓度在0.1～1.0 mg/mL范围内,其与清除率呈正相关.明日叶中黄酮类化合物对羟基自由基有较强清除效力,作为天然抗氧化产品开发具有一定价值.     

Antioxidant activity of Ixora parviflora in a cell/cell-free system and in UV-exposed human fibroblasts

Polyphenols and flavonoids possess a variety of biological activities including antioxidant and anti-tumor activities. Ixora parviflora is a member of the flavonoid-rich Rubiaceae family of flowering plants and used as folk medicine in India. The aim of this study was to investigate the antioxidant activity of Ixora parviflora extract (IPE) in a cell-free system and erythrocytes, and the ability of IPE to inhibit reactive oxygen species (ROS) generation in human fibroblasts (Hs68) after ultraviolet (UV) exposure. Various in vitro antioxidant assays were employed in this study. The extraction yield of IPE was 17.4 ± 3.9%, the total phenolic content of IPE was 26.2 μg gallic acid equivalent (GAE)/mg leaves dry weight and the total flavonoids content was 54.2 ± 4.4 μg quercetin equvalent (QE)/mg extract. The content of chlorogenic acid was 9.7 ± 1.2 mg/g extract. IPE at 1000 μg/mL exhibited a reducing capacity of 90.5 ± 0.6%, a 1,1-diphenyl-2-picrylhydrazy (DPPH) radical scavenging activity of 96.0 ± 0.4%, a ferrous chelating activity of 72.2 ± 3.5%, a hydroxyl radical scavenging activity of 96.8 ± 1.4%, and a hydrogen peroxide scavenging activity of 99.5 ± 3.3%. IPE at 500 μg/mL also possessed inhibitory activity against 2,2'-azobis (2-methylpropionamidine) dihydrochloride (AAPH)-induced hemolysis of erythrocytes (89.4 ± 1.8%) and resulted in a 52.9% reduction in ROS generation in UV-exposed fibroblasts. According to our findings, IPE is a potent antioxidant and a potential anti-photoaging agent.     

超声提取-反相高效液相色谱法测定新疆大蓟中绿原酸和芦丁的含量

以黄酮提取量为指标,选用正交实验对新疆大蓟总黄酮的超声提取工艺进行优化,结果表明最佳提取条件为:超声电流250mA,料液比1∶30(g/mL),时间40min,总黄酮提取量为2.89mg/g。同时建立了高效液相色谱法测定新疆大蓟中绿原酸及芦丁含量的方法。采用SinoChrom ODS-BP色谱柱,甲醇-1%冰乙酸为流动相,检测波长为340nm,流速为0.9mL.min-1。方法测定绿原酸的线性范围为1.675～16.75μg/mL,相关系数R=0.99995;测定芦丁的线性范围为2～20μg/mL,相关系数R=0.9999,回收率分别为99.45%、99.65%。该法简单准确,适用于新疆大蓟中绿原酸及芦丁的定量分析。     

Optimization of Enzyme-Assisted Extraction and Purification of Flavonoids from Pinus koraiensis Nut-Coated Film and Antioxidant Activity Evaluation

Pinus koraiensis nut-coated film is a kind of by-product of nut processing, which has been shown to contain flavonoids, polyphenols, and other substances that can be used to produce natural antioxidant extracts. In this study, response surface methodology (RSM) was used to optimize the extraction process of flavonoids of P. koraiensis nut-coated film (PNF), and macroporous resin HPD600 was used to purify PNF (P-PNF). Its antioxidant activity was examined by DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging capacity, oxygen free radical absorption capacity (ORAC), total oxygen radical capture (TRAP), and iron ion reduction capacity. Under the ideal extraction conditions comprising a cellulase dosage of 90 U/g, a material/liquid ratio of 1:20 (g/mL), and an extraction time of 2 h, the PNF yield was 3.37%. Purification conditions were sample concentration of 2.0 mg/mL, pH of 5, water washing volume of 3 bed volume (BV), eluent ethanol concentration of 50%, and volume of 2 BV. The P-PNF recovery was 84.32%, and purity increased from 33.80% to 61.70%. Additionally, P-PNF showed increased antioxidant activity compared to PNF. Cumulatively, this study obtained the optimal values for the process parameters in order to achieve the maximum rates of extraction of PNF for economically optimal production at an industrial scale.     

Phenolic,flavonoid contents,anticholinesterase and antioxidant evaluation of Iris germanica var; florentina

This study was designed to investigate antioxidant and anticholinesterase potential of Iris germanica var; florentina. Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory potential of plant samples were investigated by Ellman’s assay. Antioxidant activity was performed using DPPH, H₂O₂ and ABTS free radical scavenging assays. Total phenolics and flavonoids contents were expressed in mg GAE/g dry weight and mg RTE/g, respectively. In AChE inhibition assay, Ig.Fl, Ig.Sp and Ig.Cf fractions exhibited highest activity with IC₅₀ values of < 0.1, 5.64 and 19 μg/mL, respectively. In BChE inhibitory assay, Ig.Fl, Ig.Sp, Ig.Cf and Ig.Cr were most active with IC₅₀ of < 0.1, < 0.1, 31 and 78 μg/mL, respectively. In DPPH assay, Ig.Fl and Ig.Cf exhibited highest inhibition of free radicals, 80.52% (IC₅₀ = 9 μg/mL) and 78.30% (IC₅₀ = 8 μg/mL), respectively. In ABTS assay Ig.Cr, Ig.Cf, Ig.Fl and Ig.Sp exhibited IC₅₀ values of < 0.1, 2, 2 and 3 μg/mL, respectively.     

Study on Extraction and Antioxidant Activity of Flavonoids from Hemerocallis fulva (Daylily) Leaves

Hemerocallis fulva is a medical and edible plant. In this study, we optimized the ultrasound-assisted extraction (UAE) process of extracting flavonoids from Hemerocallis fulva leaves by single-factor experiments and response surface methodology (RSM). The optimum extraction conditions generating the maximal total flavonoids content was as follows: 70.6% ethanol concentration; 43.9:1 mL/g solvent to sample ratio; 61.7 °C extraction temperature. Under the optimized extraction conditions, the total flavonoid content (TFC) in eight Hemerocallis fulva varieties were determined, and H. fulva (L.) L. var. kwanso Regel had the highest TFC. The cytotoxicity of the extract was studied using the Cell Counting Kit-8 (CCK-8 assay). When the concentration was less than 1.25 mg/mL, the extract had no significant cytotoxicity to HaCaT cells. The antioxidant activity was measured via chemical antioxidant activity methods in vitro and via cellular antioxidant activity methods. The results indicated that the extract had a strong ABTS and •OH radical scavenging activity. Additionally, the extract had an excellent protective effect against H₂O₂-induced oxidative damage at a concentration of 1.25 mg/mL, which could effectively reduce the level of ROS to 106.681 ± 9.733% (p < 0.001), compared with the 163.995 ± 6.308% of the H₂O₂ group. We identified five flavonoids in the extracts using high-performance liquid chromatography (HPLC). Infrared spectroscopy indicated that the extract contained the structure of flavonoids. The results showed that the extract of Hemerocallis fulva leaves had excellent biocompatibility and antioxidant activity, and could be used as a cheap and potential source of antioxidants in the food, cosmetics, and medicine industries.     

Rapid screening,separation, and detection of hydroxyl radical scavengers from total flavonoids of Ginkgo biloba leaves by chromatography combined with molecular devices

Hydroxyl radicals are the most reactive free radical of human body, a strong contributor to tissue damage. In this study, liquid chromatography coupled to electrospray ionization mass spectrometry was applied to screen and identify hydroxyl radical scavengers from the total flavonoids of Ginkgo biloba leaves, and high‐performance counter current chromatography was used to separate and isolate the active compounds. Furthermore, molecular devices were used to determine hydroxyl radical scavenging activities of the obtained hydroxyl radical scavengers and other flavonoids from G. biloba leaves. As a result, six compounds were screened as hydroxyl radical scavengers, but only three flavonoids, namely, rutin, cosmos glycosides and apigenin‐7‐O‐Glu‐4’‐O‐Rha, were isolated successfully from total flavonoids by high‐performance counter current chromatography. The purities of the three obtained compounds were over 90%, respectively, as determined by liquid chromatography. Molecular devices with 96‐well microplates evaluation indicated that the 50% scavenging concentration values of screened compounds were lower than that of other flavonoids, they performed greater hydroxyl radical scavenging activity, and the evaluation effects were consistent with the liquid chromatography with mass spectrometry screening results. Therefore, chromatography combined with molecular devices is a feasible and an efficient method for systematic screening, identification, isolation, and evaluation of bioactive components in mixture of botanical medicines.     

流动注射-抑制化学发光法测定银杏叶中总黄酮含量

提出了流动注射-抑制化学发光测定银杏叶中的总黄酮含量的分析方法.它是基于银杏叶中物质黄酮类具有还原性,在碱性条件下还原H_2O_2,抑制鲁米诺-H_2O_2-KIO_4体系的化学发光,其抑制程度的大小与总黄酮的含量成线性关系.方法的线性范围为1.5～30μg/mL,检出限为0.03μg/mL,相对标准偏差(RSD)为1.2%,采样频率为240次/h,回收率为101%～104%.