Phlorizin alleviates deltamethrin-induced oxidative stress in brine shrimp Artemia

Deltamethrin(DEL),a commonly used pyrethroid pesticide,results in higher reactive oxygen species(ROS)levels in aquatic animals,which consequently unbalance the redox state.Phlorizin(PHL)is a flavonoid and a natural product promising to prevent or reduce pesticide-induced oxidative stress.Artemia is a micro-crustacean widely used in marine hatcheries and an experimental aquatic organism for environmental toxicology research.This research aimed to evaluate the toxicity of DEL on Artemia and the antioxidative effect of PHL against the toxicity.Results show that 0.08-mg/mL PHL exerted its antioxidative effects on hatching percentage of the cysts in 24-h incubation and on body length and survival rate of Artemia in 12-d culture.After 12-d culture,12-,24-,and 36-h DEL exposure showed significant drops in SOD,CAT,and GSH-Px enzyme activities,and significant increases in ROS and malondialdehyde(MDA)levels in Artemia(P<0.05).On the contrary,0.08-mg/mL PHL application improved the enzyme activities and decreased the ROS and MDA levels(P<0.05).Moreover,0.08-mg/mL PHL significantly increased mRNA expression levels of Cu/Zn SOD,CAT,GST,HO-1,NQO1,and Nrf2,and decreased mRNA expression level of Keap1 in the DEL-exposed Artemia(P<0.05).Therefore,DEL is toxic to Artemia,while PHL alleviates DEL-induced oxidative damage by possibly regulating the Nrf2signaling pathway.This study provided a theoretical basis for PHL to reduce pesticide-induced toxicity in aquatic animals.

Evaluation of the binding affinity and antioxidant activity of phlorizin to pepsin and trypsin

Phlorizin(PHL)is a natural compound with strong antioxidant properties mainly found in apples.In this paper,the interaction mechanism of PHL with pepsin and trypsin was comparatively evaluated by computer simulation,fluorescence spectra,circular dichroism(CD),and Fourier transform infrared(FT-IR)spectra at a molecular level.Fluorescence spectra showed that PHL quenches the pepsin/trypsin by static quenching.Thermodynamic parameters indicated that PHL binds to pepsin mainly through hydrogen bonds and van der Waals forces,and that of trypsin was electrostatic forces.The ground state complexes PHL and protease have a moderate affinity of 105 L/mol PHL binds more strongly to trypsin than to pepsin.CD and FT-IR spectra results showed that pepsin/trypsin decreased theβ-sheet content and slightly changed its secondary structure upon PHL.These experimental results are mutually verified with the predicted computer-aid simulation results.Upon PHL and trypsin binding,the antioxidant capacity of PHL was elevated.Nevertheless,the antioxidant capacity of PHL was decreased after binding to pepsin.This work elucidates the binding of PHL binding mechanisms to pepsin/trypsin and provides useful information for the digestion of PHL to improve the application of PHL in food processing.

Effect of phlorizin on metabolic abnormalities in Spontaneously Diabetic Torii (SDT) rats

The Spontaneously Diabetic Torii (SDT) rat is a novel model for nonobese type 2 diabetes. In this study we investigated the glycolipid metabolic changes with phlorizin-treatment, which inhibits intestinal glucose uptake and renal glucose reabsorption, in male SDT rats. Phlorizin (100 mg/kg, b.i.d., s.c.) was administered for 4 weeks to SDT rats from 20 to 24 weeks of age. As a result, phlorizin reduced the development of hyperglycemia and decreased the hemoglobin A1c (HbA1c) levels. In the liver, phlorizin increased mRNA levels of glucokinase, the enzymes related with the glycogen cascade and the proteins associated with lipid metabolism. In conclusion, chronic administration of phlorizin in SDT rats produced a good glycemic control and an improvement in liver function.

Immune Reactive Ezrin Surface Area Increases in Glomerular Podocytes of STZ Diabetic Rats Precede Their Detachment, Is Prevented by Phlorizin But Not by Insulin

Glomerular tuft immune reactive Ezrin surface area (EzA) and fraction of EzA to total glomerular tuft area significantly increased, indicating podocyte growth, rounding and altered cytoskeletal interactions at 1 week of STZ diabetes. Podocyte number per glomerulus (WT1+ nuclei) did not change indicating no detachment, but density decreased due to tuft hypertrophy. Treatment with PLZ or Insulin for one week, prevented increase in proteinuria and hyperglycemia but not the decrease in podocyte density. PLZ but not Insulin prevented increase in ezrin positive area in glomeruli and per podocyte. In podocytes in culture neither 25 mM glucose with or without PLZ (2.5 or 25 uM) altered Ezrin expression measured in western blots. In summary, the Ezrin positive glomerular surface area increase seen after 1 week of STZ diabetes, reflects altered podocyte morphology and cytoskeletal interactions, prevented by PLZ but not by insulin. Ezrin area increase preceded podocyte detachment and in podocytes in culture is not associated with increases in podocyte Ezrin protein expression. It is a likely precursor of shape changes in podocytes and of alterd interactions with basement membrane that contribute to detachment and thickening. Glomerular capillary tuft hypertrophy and reduced podocyte density persisted despite PLZ or insulin treatments, independently of levels of glycemia and of proteinuria.

不同浓度根皮苷对苹果砧木根系生长及生理特性的影响

本试验以苹果砧木组培苗为试材,设置0(CK)、0.1、0.5、1.0 mmol/L和1.5 mmol/L共5个根皮苷浓度处理,研究砂培条件下各处理对苹果砧木组培苗生长、根系构型及根系生理生化特性的影响。结果表明,与CK相比,外施不同浓度根皮苷均抑制苹果砧木组培苗根系生长,显著降低组培苗鲜重和干重,均显著降低根长、根表面积、根体积、根平均直径和株高净增长量(0.1 mmol/L浓度处理除外),且随着根皮苷浓度升高而降幅增大。外源根皮苷胁迫导致苹果砧木组培苗根系超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)活性降低,与根系根皮苷代谢相关的查尔酮合酶、查尔酮异构酶及根皮苷水解酶活性显著降低,丙二醛(MDA)、超氧阴离子、脯氨酸含量显著升高,可溶性蛋白含量显著降低。综之,随着根皮苷浓度增加,苹果砧木组培苗根系抗氧化保护性酶活性和可溶性蛋白含量降低,MDA、超氧阴离子含量及脯氨酸含量升高,根系生长受到抑制,从而抑制苹果砧木组培苗生长。

Effects of phlorizin on vascular complications in diabetes db/db mice

Background Diabetic macrovascular complications are important causes of cardiovascular and cerebrovascular diseases and also one of the major causes of morbidity and mortality in patients with type 2 diabetes mellitus （T2DM）. Phlorizin has been reported to be effective in reducing the blood glucose level in diabetic mellitus, while little is known about its effects on vascular complications. This study aimed to observe the effects of phlorizin on the aorta of diabetes db/db mice and explore its mechanism. Methods Diabetic db/db mice （n=16） and age-matched db/m mice （n=8） were divided into three groups： normal control group （CC group, db/m mice, n=8）, untreated diabetic group （DM group, db/db mice, n=8） and diabetic group treated by phlorizin （DMT group, db/db mice, n=8）. Phlorizin （20 mg/kg body weight） was given in normal saline solution intragastrically for 10 weeks. Animals were weighed weekly. At the 10th weekend, all mice were fasted overnight and then sacrificed. Fasting blood was collected, and the aortas were dissected. The blood samples were analyzed for fasting blood glucose （FBG）, serum advanced glycation end products （AGEs）, malondialdehyde （MDA） and superoxide dismutase （SOD） activity, the aortic ultrastructure was studied. Results The weight and serum concentration of FBG, AGEs, and MDA in the DM group were higher than that in the CC group （P 〈0.01 ）, and they were significantly lower in the DMT group （P 〈0.05）. Serum SOD activity was lower than that in the CC group （P 〈0.01）, and it is significantly higher in the DMT group （P 〈0.05）. The severity of aorta damage in the DMT group was less than that in the DM group. Conclusions Phlorizin protected the db/db mice from diabetic macrovascular complications, attributed to the decreasing of blood glucose and AGEs level, and its antioxidant potential. This study may provide a new natural medicine for treating diabetic macrovascular complications.

5种天然多酚类化合物抗氧化活性的比较

文中以VC、BHT和EDTA为对照,通过总抗氧化能力、DPPH自由基清除活性、ABTS自由基清除活性、铁离子还原能力和金属离子螯合能力等5种体系对咖啡酸、芦丁、白藜芦醇、根皮素和根皮苷等天然多酚类化合物的抗氧化活性进行了综合评价。结果表明:咖啡酸的总抗氧化能力、DPPH自由基清除能力[IC50(0.023±0.004)mg/mL]和铁离子还原能力均为最高,在浓度为0.1 mg/mL时总抗氧化能力达到(1.873±0.022)mg VC eq/mg;白藜芦醇的ABTS自由基清除能力最强[IC50(0.056±0.006)mg/mL],极显著高于阳性对照BHT和VC(P<0.01);芦丁的金属离子螯合能力最强,白藜芦醇次之,其他多酚类化合物不具备金属离子螯合能力。咖啡酸、芦丁、白藜芦醇、根皮素和根皮苷均具有较强的抗氧化活性,是潜在的天然抗氧化剂。

短果杜鹃组培快繁及其种质试管保存培养基的筛选

以短果杜鹃新生嫩芽为外植体,应用均匀设计法筛选其基部直接再生芽苗、生根及其试管苗保存培养基,结果表明,最适合的基部直接再生芽苗诱导培养基为:DR+2-ip2.75mg.L-1+IAA0.07mg.L-1;生根培养基:1/2MS(大量元素)+IAA0.1mg.L-1+NAA0.05mg.L-1+Kt0.2mg.L-1;试管苗保存培养基:1/5MS+根皮苷5.0mg.L-1。短果杜鹃组织培养的不同阶段需要不同的培养基,常温条件下,利用"低营养延缓生长"的方法在试管内保存其种质。

光谱法研究根皮苷与人血清白蛋白相互作用

在模拟生理条件下,采用荧光光谱、紫外光谱和同步荧光光谱法研究了根皮苷(Phlorizin)与人血清白蛋白(HSA)的相互作用。结果表明:根皮苷能使HSA发生内源荧光猝灭,属静态猝灭。在293、303 K和313 K下,根皮苷与HSA的结合常数分别为3.163 5×105、1.774 8×105、1.193 5×105L.mol-1,结合位点数n近似为1;热力学分析表明根皮苷与HSA间的结合力为氢键及范德华力;根据Frster非辐射能量转移理论求得二者结合距离为3.97 nm;同步荧光光谱表明根皮苷主要与HSA中的色氨酸残基发生相互作用,改变色氨酸周围的局部构象;金属离子的介入会影响根皮苷与HSA的结合能力。

植物根际放线菌分离方法初探及根皮苷降解活性分析

【目的】研究不同培养基及不同土样处理方法对植物根际放线菌分离效果的影响,并从中挖掘具有开发潜力的高活性根皮苷降解放线菌株,为苹果连作障碍机制研究及微生物修复提供理论和实践指导。【方法】从吉林、河北、山东、陕西、甘肃、河南、湖北、四川、重庆、上海、江西、安徽、浙江等13个省(市)采集植物根际土样40份,自然风干处理后分别经10mmol/L的MOPS溶液及CaCO3预处理,用涂布平板法在7种不同的培养基中进行放线菌的分离培养。将分离获得的菌株点种于根皮苷筛选培养基进行活性检测。【结果】在7种培养基B、MSC、MG、MT、Chi、HSG、HV中分别分离获得100、78、108、64、79、113和88株放线菌。土样用MOPS溶液处理后所分离得到的放线菌总数明显高于用CaCO3预处理所得。分离所得630株菌株中有29株对根皮苷有降解活性。对其中41株高活性菌株及形态特殊菌株进行16SrRNA基因测序分析,发现分属于14个不同的放线菌属。【结论】7种分离培养基中HSG培养基的分离效果最好,MG培养基次之,MT培养基最差。土样处理中用MOPS溶液处理的方法优于用CaCO3预处理。根际放线菌株中有一定数量具有降解根皮苷能力的菌株。根际放线菌资源具有丰富的生物多样性。

根皮苷对高脂膳食喂饲仓鼠氧化损伤保护机制的研究

以饲喂高脂膳食的仓鼠为动物模型,研究根皮苷在喂饲高脂膳食仓鼠体内的抗氧化活性。实验动物随机分为对照组和3个实验组(3、6、9 g/kg根皮苷),6周后测定仓鼠血清、心脏、肾脏及肝脏中抗氧化酶超氧化物歧化酶(superoxide dismutase,SOD)、谷胱甘肽过氧化物酶(glutathion peroxidase,GSH-Px)、过氧化氢酶(catalase,CAT)和总抗氧化力(total antioxidant capacity,T-AOC)的活力、丙二醛(malonaldehyde,MDA)含量及采用荧光定量聚合酶链式反应(real time-polymerase chain reaction,RT-PCR)法检测肝脏中抗氧化相关基因mRNA表达水平。抗氧化酶活力测定结果显示,给予根皮苷后血清、心脏、肾脏及肝脏中SOD、GSH-Px、CAT酶活力均有升高,氧化产物MDA含量显著降低。RT-PCR结果显示,3个不同剂量根皮苷实验组CuZn-SOD、Mn-SOD、GSH-Px、血红素加氧酶1(heme oxygenase 1,HO-1)、核因子E2相关因子2(nuclear factor erythroid-2 related factor 2,Nrf2)mRNA表达水平与对照组相比均极显著升高(P<0.01);给予6、9 g/kg根皮苷,CAT基因表达水平较对照组极显著升高(P<0.01)。因此,根皮苷对高脂膳食喂饲仓鼠氧化应激损伤的保护作用可能是通过激活抗氧化基因Nrf2表达,并上调抗氧化基因SOD、GSH-Px、CAT、HO-1 mRNA表达水平,进而增加抗氧化酶的活力实现的。

根皮苷通过下调小肠NPC1L1和HMG-CoA还原酶表达降低血液胆固醇水平

以高脂高胆固醇膳食喂饲仓鼠为动物模型,研究根皮苷对仓鼠血脂水平及小肠胆固醇代谢相关基因的调控影响。36只实验动物随机分成对照组和3个不同剂量根皮苷干预组(3、6、9 g/kg),测定血清中总胆固醇(total cholesterol,TC)、甘油三酯(triglyceride,TG)及高密度脂蛋白胆固醇(high density lipoprotein cholesterol,HDL-C)水平,气相色谱法检测肝脏中胆固醇含量及粪便固醇排泄量,实时定量荧光聚合酶链式反应(real-time polymerase chain reaction,Real-Time PCR)分析小肠胆固醇合成、吸收、转化及排泄基因的表达水平。血脂测定结果显示,血清TC、TG水平随给予根皮苷添加量增加而降低,且高剂量组具有显著性(P<0.05或P<0.01);6、9 g/kg根皮苷剂量组,血清中HDL-C水平极显著升高(P<0.01)。气相色谱检测结果显示,根皮苷剂量组仓鼠肝脏中胆固醇含量较对照组随根皮苷剂量增加而降低,且6、9 g/kg组具有极显著差异(P<0.01);粪便中总中性固醇排泄量与根皮苷给予量正相关,且都具有显著性差异(P<0.05或P<0.01),总酸性固醇排泄量根皮苷剂量组相比对照组增加,且6 g/kg组显著增加(P<0.05),9 g/kg组极显著增加(P<0.01)。Real-Time PCR检测结果显示,根皮苷剂量组小肠中胆固醇合成限速酶HMG-CoA还原酶、小肠胆固醇吸收关键蛋白NPC1L1、胆固醇酯化酶ACAT2、微粒体转运蛋白MTP的mRNA表达水平较对照组显著降低(P<0.05或P<0.01);给予根皮苷后,肠道中促进胆固醇向外排泄基因ABCG5/8表达水平显著升高(P<0.05或P<0.01)。因此,根皮苷对高脂高胆固醇膳食饲喂仓鼠胆固醇代谢平衡的调节可能是通过对小肠胆固醇吸收、转化等基因表达的抑制,上调胆固醇排泄基因的表达实现的。

根皮苷对雌性果蝇寿命的影响及其作用机制

目的:研究根皮苷(phloridzin)对雌性果蝇寿命的影响并探究其延缓果蝇衰老的分子机制。方法:在果蝇寿命实验中,采用2日龄雌性果蝇作为研究对象,在其培养基中分别加入0、0.5、2.0、5.0 mg/m L根皮苷进行培养,直至果蝇全部死亡,计算果蝇的平均寿命和最长寿命;通过白草枯和双氧水急性实验测定根皮苷对果蝇氧化应激损伤的影响;试剂盒法测定果蝇体内抗氧化酶活力;实时荧光定量聚合酶链式反应(real-time polymerase chain reaction,real-time PCR)检测抗氧化基因表达水平。结果:2.0、5.0 mg/m L根皮苷可以显著延长果蝇的平均寿命和最长寿命(P<0.05);根皮苷能够对百草枯和双氧水造成的雌性果蝇氧化应激损伤起到有效保护作用;添加5.0 mg/m L根皮苷可以极显著提高Cu/Zn-超氧化物歧化酶(superoxide dismutase,SOD)和过氧化氢酶(catalase,CAT)活力(P<0.01),极显著降低丙二醛(malonaldehyde,MDA)含量(P<0.01);添加5.0 mg/m L根皮苷可以极显著上调Cu/Zn-SOD及CAT m RNA表达水平(P<0.01)。结论:根皮苷能够延缓雌性果蝇衰老,且具有抗氧化作用,这可能与其调控抗氧化基因的表达水平相关。

苹果树腐烂病菌发酵液中根皮苷主要降解成分的分析

【目的】明确苹果树腐烂病菌致病过程中对降解根皮苷发挥主要作用的物质,为揭示该病原菌的致病机理提供理论依据。【方法】以根皮苷为碳源发酵培养苹果树腐烂病菌10d后,测量菌丝干质量及病原菌对根皮苷的降解率;以淀粉为碳源获取病原菌30d的无菌发酵液,进一步分离发酵液中的有机酸和蛋白类物质。用HPLC法分别检测病原菌、淀粉无菌发酵液、有机酸类、蛋白类物质及发酵剩余液对根皮苷的降解作用。【结果】以根皮苷为碳源发酵培养病原菌10d后,每100mL发酵液中菌丝干质量为48mg,病原菌对根皮苷的降解率为92.46%,发酵液对根皮苷的降解率为99.88%。1mg/mL蛋白类物质与根皮苷作用5d的降解率为90.4%,有机酸类对根皮苷的降解率仅为1.6%,发酵剩余液对根皮苷的降解率为5.9%。根皮苷的降解率随蛋白液质量浓度的增加和作用时间的延长而升高。【结论】在根皮苷降解过程中起主要作用的物质为苹果树腐烂病菌代谢产生的蛋白,其可能为细胞壁降解酶类。

超声波辅助提取苹果叶多酚及其抗氧化性研究

研究了超声波辅助提取苹果叶多酚的最佳工艺及其抗氧化性。结果表明,超声波辅助提取苹果叶多酚的最佳条件为:乙醇体积分数20%,液料比40∶1(mL∶g),提取温度75℃,提取时间20 min,超声波功率600 W,提取两次,在此条件下多酚的得率为4.22%;苹果叶提取物有较好的抗氧化活性,对.OH和O2-.的清除能力比茶多酚强;HPLC分析表明苹果叶中根皮苷的含量为2.45%,苹果叶提取物中根皮苷的含量为6.18%,占提取物中多酚量的61.36%。

根皮苷的应用及分离纯化研究进展

根皮苷具有降低血糖、抗氧化等作用,目前已被应用于医药、化妆品、食品、植物组织培养等领域.对根皮苷的应用及目前用于分离纯化根皮苷的方法进行了综述,以期对根皮苷的开发利用提供帮助.

紫变青霉生物转化根皮苷制备根皮素的研究

为了开发一种生物转化根皮苷生成根皮素的工艺,首先从苹果皮富集培养物中分离筛选微生物菌株,进而优化产酶培养基主要组成、pH和转化条件,提高根皮素的转化得率.结果从富集培养物中筛选到一株紫变青霉(Penicillium purpurescens)QL-9204菌株,经产酶培养制备的粗酶液能转化根皮苷为根皮素;在产酶培养基组成为蔗糖2g/L,(NH4)2SO46g/L,NaCl 5g/L,KH2PO45g/L,MgSO41g/L,MnSO41g/L,pH=6时,QL-9204菌株经种子扩培后接种产酶培养基,于30℃,200r/min条件下培养5d,滤除菌体的粗酶液中加入80mg/L的根皮苷,于30℃,250r/min转化14h后,根皮素的转化得率可达90.3%.利用紫变青霉菌QL-9204发酵制备的粗酶液转化根皮苷制备根皮素,具有方法简单、转化得率高和成本低的优势,有潜在的工业化应用价值.

苹果叶多酚的纯化及其抗氧化性研究

对苹果叶多酚的纯化及其抗氧化性进行研究。采用大孔吸附树脂对苹果叶多酚进行初步纯化、Sephadex LH-20进一步精制,并进行高效液相分析。结果表明X-5树脂对苹果叶多酚有较好的吸附解吸效果,X-5树脂纯化苹果叶多酚的条件为上柱液质量浓度3.658mg/mL、上柱液pH3、吸附流速2.0mL/min;以体积分数40%乙醇为解吸剂,洗脱流速1.0mL/min,洗脱体积4BV。在此条件下纯化后的苹果叶多酚的纯度从10.07%提高到38.55%;X-5树脂对苹果叶多酚的吸附为放热过程,吸附过程符合Langmuir等温吸附模型和Freundlich等温吸附模型。经过X-5树脂纯化后的苹果叶多酚对DPPH自由基和NO-2·的清除能力有所增强;苹果叶中根皮苷的含量为2.45%,粗提物经过X-5树脂纯化和Sephadex LH-20精制后得到的苹果叶多酚精制品中根皮苷的含量为47.61%,占制品中总多酚的91.51%。

根皮素抑酪氨酸酶活性研究

根皮素从苹果皮中提取分离 ,测定其对酪氨酸酶的活性抑制率。浓度为 0 .3 %时根皮素的抑制率可达 98.2 % ,其IC50 为 0 .0 5 % ,结果优于曲酸和熊果苷。与曲酸和熊果苷复配 ,可使抑制率达到 1 0 0 %。

三种海棠黄酮类成分分析

目的:对三种海棠进行黄酮类成分分析,为完善湖北海棠的药用标准及资源利用提供依据。方法:采用薄层色谱(TLC)法对三种海棠进行定性鉴别;采用紫外分光光度法测定三种海棠中总黄酮的含量;采用高效液相色谱(HPLC)法测定根皮苷的含量。结果:聚酰胺TLC分离效果良好,斑点清晰。6批湖北海棠中总黄酮含量范围为19.65～20.97 mg/g,变叶海棠中总黄酮含量范围为10.71～12.13 mg/g,平邑甜茶中总黄酮含量范围为3.49～3.67 mg/g;湖北海棠中根皮苷含量为15.51 mg/g,变叶海棠中根皮苷含量为14.66 mg/g,平邑甜茶中根皮苷含量为2.05 mg/g。结论:所建方法有助于进行不同来源海棠的黄酮类成分定性及定量研究,为进一步完善湖北海棠的药用标准及规范湖北海棠的品种应用奠定基础。