Tyrosinase inhibitory effects and inhibition mechanisms of nobiletin and hesperidin from citrus peel crude extracts

The inhibitory effects of nobiletin and hesperidin from citrus peel crude extracts on tyrosinase diphenolase activity are evaluated. IC₅₀ of nobiletin and hesperidin is 1.49 mM and 16.08 mM, respectively and their inhibition mechanism is competitive type with K_i = 2.82 mM and noncompetitive with K_i = 9.16 mM, respectively. Crude extracts from citrus peel (C. unshiu Marc.) were extracted with 95% ethanol and fractionated by petroleum ether (PCPE). The ethanol phase (ECPE) was further desorbed from macroporous adsorption resin (FGRE). Their IC₅₀ values were 8.09 mg/mL, 7.53 mg/mL and 4.80 mg/mL, respectively. Their inhibition on melanogenesis in B16 mouse melanoma cells was also evaluated. FGRE showed a significant inhibition (42.5% at 31.25 μg/mL, p < 0.01) while hesperidin showed almost no inhibition. Nobiletin and PCPE give efficacious antiproliferation effects on B16 mouse melanoma cell with IC₅₀ values 88.6 μM and 62.96 μg/mL, respectively, by the MTT test. Hesperidin and other crude extracts showed very low cytotoxity to the B16 cell.     

ATGL调节细胞内脂质代谢和代谢相关疾病

脂肪组织甘油三酯水解酶(adipose triglyceride lipase, ATGL)是一种催化甘油三酯第一步水解的重要脂肪酶,在机体能量代谢调节中发挥重要作用.本文介绍了ATGL的基因和蛋白质结构,并详细综述了ATGL的功能调控和与其相关联疾病的研究进展,最后通过与激素敏感脂肪酶(HSL)比较,对ATGL的特征进行总结.     

Hesperidin,a citrus flavonoid,increases the bioavailability of micronutrients of Gallus domesticus (chicken) eggshell: in vitro study

The consumption of citrus flavonoid, hesperidin may inhibit the bone loss. The purpose of this study was to evaluate the effect of hesperidin on the bioavailability of Ca, a probable reason to prevent bone loss. Citrus flavonoid (hesperidin) in combination with citric acid and ascorbic acid was scrutinized to estimate the bioavailability of micronutrients from chicken egg shells using in vitro method. Effect of citric acid, ascorbic acid and hesperidin on the bioavailability of minerals (Zn, Fe) and macro elements (Ca, Mg, P) was evaluated and the amounts required to get maximum bioavailability were concluded. The highest bioavailability of Ca, Mg, P, Fe and Zn was 89.25 ± 2.13, 92.28 ± 1.87, 40.32 ± 3.09, 32.81 ± 1.24 and 46.19 ± 0.83%, respectively after the addition of 3 g of citric acid, 100 mg of ascorbic acid and 4 mg of hesperidin per gram of chicken eggshell powder. Citric acid greatly affects the bioavailability of Ca, Mg, P, and Zn, whereas addition of ascorbic acid enhances the bioavailability of Fe, and hesperidin boosts the bioavailability (p < 0.05) of all micronutrients of the chicken eggshells.     

Crystallization and preliminary crystallographic studies of the precursor and mature forms of a neutral lipase from the fungus Rhizopus delemar

A neutral lipase from the filamentous fungus Rhizopus delemar has been crystallized in both its proenzyme and mature forms. Although the latter crystallizes readily and produces a variety of crystal forms, only one was found to be suitable for X-ray studies. It is monoclinic (C2, a = 92.8 Å, b = 128.9 Å, c = 78.3 Å, β = 135.8) with two molecules in the asymmetric unit related by a noncrystallographic diad. The prolipase crystals are orthorhombic (P2₁2₁2₁, with a = 79.8 Å, b = 115.2 Å, c = 73.0 Å) and also contain a pair of molecules in the asymmetric unit. Initial results of molecular replacement calculations using the refined coordinates of the related lipase from Rhizomucor miehei identified the correct orientations and positions of the protein molecules in the unit cells of crystals of both proenzyme and the mature form. © 1994 John Wiley & Sons, Inc.     

温州蜜柑果实发育进程中光合产物运输分配及糖积累特性

测定了温州蜜柑果实发育进程中糖的积累动态,并用14C示踪方法研究了果实发育不同阶段源叶的光合产物在果实中的运输分配特性.结果表明(1)果实组织中的糖含量随果实发育持续上升,果皮以积累己糖为主而其它组织则以积累蔗糖为主或略多.(2)在果实完熟之前,汁囊分配到的光合产物均占果实的50%以上,为果实中的光合产物分配中心;而到完熟阶段分配到汁囊的14C光合产物与其它组织接近,不再有明显的分配中心.(3)汁囊中的14C放射性比活度随着果实发育呈下降趋势,尤其在后期下降极为明显.(4)维管束/囊瓣表皮和囊瓣表皮/汁囊的14C放射性比活度的比率都随果实发育进程而增大,但后者的比率要大大高于前者,表明韧皮部后运输是光合产物进入汁囊的限速阶段.(5)输入到汁囊的14C光合产物主要以蔗糖形式存在,说明光合产物是以蔗糖为主体输入汁囊参与糖积累和代谢的.     

温州蜜柑叶片气体交换和叶绿素荧光对低温的响应

研究了低温对温州蜜柑叶片气体交换和叶绿素荧光的影响 ,结果表明 :(1) 8℃低温处理 18h对气体交换和叶绿素荧光影响不大。 (2 ) 2℃低温处理 15h后 ,净光合速率 (Pn)、气孔导度 (Gs)、羧化效率 (CE)下降 ,胞间CO2 浓度 (Ci)升高 ,表观量子效率 (AQY)和叶绿素荧光参数F0 及Fv/Fm没有显著变化。 (3)室外自然低温处理 2和 7d ,Pn、Gs、CE、饱和CO2 光合速率、AQY及Fv/Fm显著下降 ,Ci及F0 显著升高 ;在 2 0℃室内 ,Fv/Fm、F0 和AQY比CE和Pn恢复快。 (4 )低温胁迫使Jf(依据叶绿素荧光参数计算所得的电子传递速率 )和Jc(依据CO2 同化测定所得的电子传递速率 )下降 ,Jf/Jc 值升高。 (5 )低温处理降低了Pn和光呼吸速率 (Pr) ,但Pr下降的速率小于Pn下降的速率     

Abscisic acid increases in non-growing and paclobutrazol-treated fruits of seedless mandarins

Clementine ( Citrus reticulata [Hort.] Ex. Tanaka cv. Oroval) is a self-incompatible mandarin with a slow rate of fruit growth and high percentage of fruit abscission. Seedless Satsuma ( Citrus unshiu [Mak] Marc. cv. Clausellina) shows natural parthenocarpy and higher fruit set. Application of 25 μg fruit⁻¹ of paclobutrazol (PP333), an inhibitor of gibberellin biosynthesis, reduced the rate of growth and accelerated fruit abscission in both varieties. In contrast, gibberellin A₃ (GA₃) stimulated fruit growth only in the self-incompatible mandarin. Clementine fruits, in the absence of pollination, showed an approximately 2-fold transient increase in the free abscisic acid (ABA) content shortly after petal fall. In Satsuma, a very small accumulation of ABA was detected. Paclobutrazol treatment induced a 3-fold increase in ABA in Satsuma fruits but did not substantially affect the pattern of ABA accumulation in Clementine. In this variety, GA₃ suppressed the ABA increase observed in untreated fruits. These effects were observed 24 h after treatment. However, in Satsuma fruits, the effect of GA₃ on the ABA content was negligible. In addition, a comparative analysis of growing and non-growing fruits of Clementine showed that ABA, on a per unit weight basis, was always higher in the non-growing fruits. Treatment with 85 μM fluridone, an inhibitor of carotenoid biosynthesis and thus indirectly of ABA, delayed fruit abscission in Clementine, but also decreased fruit growth. Collectively, these observations indicate a relationship between high ABA content and a reduced rate of fruit growth and an acceleration of fruit abscission.     

Effects of Citrus unshiu Powder on the Cytokine Balance in Peripheral Blood Mononuclear Cells of Patients with Seasonal Allergic Rhinitis to Pollen

We evaluated the effects of a 50% methanol extract of Citrus unshiu powder (MEC) on cytokines in peripheral blood mononuclear cells (PBMCs) obtained from patients with seasonal allergic rhinitis to cedar pollen. The levels of cytokines, such as TNF-α, IFN-γ, IL-2, IL-4, IL-5, IL-10, IL-12 (p70), IL-13, and GM-CSF, produced by pollen-stimulated PBMC were measured. We found that MEC suppressed pollen-induced TNF-α release and increased IFN-γ release from PBMCs. The results suggest that Citrus unshiu powder has an immunomodulatory effect in vitro and that its use could improve seasonal allergic rhinitis symptoms.     

枳实中辛弗林和橙皮苷的联合提取工艺研究

以辛弗林和橙皮苷的收率及纯度为指标,研究了含辛弗林的枳实提取物及橙皮苷的综合制备工艺.先采用酸液渗漉法提取枳实中的辛弗林,再采用碱提酸沉法提取原料中的橙皮苷,考察盐酸浓度及用量、流速等因素的影响.辛弗林的最佳提取工艺为:药材加5倍水浸泡过夜,渗漉盐酸液的浓度为0.02 mol/mL,渗漉料液比为1:4,以5 mL/min的流速渗漉;碱提酸沉法提取橙皮苷的优化工艺为:于提取辛弗林之后的药渣中加入65%乙醇,于60℃搅拌条件下加碱调节pH 13,过滤后的滤液以稀盐酸调节pH 5,过滤后再用碱回调pH7,放置过夜.橙皮苷的收率为11.3%,HPLC测定橙皮苷的纯度为86.7%.     

脂蛋白酯酶在动脉粥样硬化发生发展中的作用

脂蛋白酯酶(lipoprotein lipase, LPL)是调节甘油三酯代谢的关键酶,在动脉粥样硬化(atherosclerosis,As)的发生发展中起重要作用。LPL产生部位的差异决定了其具有促As作用还是抗As作用。其次,不同因素对LPL的调控也会使LPL对As产生相反的作用效果。本文综述了LPL在As发生发展中的作用机制以及不同因素对LPL的调控机制,对于As的防治具有重要意义。     

Low temperature influence on flowering in Citrus. The separation of inductive and bud dormancy releasing effects

The flowering response of Owari Satsuma mandarin ( Citrus unshiu Marc) to low temperature treatments has been determined using potted trees and in vitro bud cultures. In potted trees the chilling treatments released bud dormancy and enhanced both sprouting and flowering, but these two responses could not be separated. However, bud cultures showed no dormancy, and a specific effect of low temperature on flower induction was demonstrated. Low temperature appears to have a dual effect, releasing bud dormancy and inducing flowering. Potential flower buds have a deeper dormancy than vegetative buds, and the first stages of flower initiation seem to occur before the winter rest period.     

Hormonal changes associated with fruit set and development in mandarins differing in their parthenocarpic ability

Satsuma [Citrus unshiu (Mak) Marc.] and Clementine [Citrus reticulata (Hort.) Ex. Tanaka, cv. Oroval] are two related species of seedless mandarins which differ in their tendency to set parthenocarpic fruits. Satsuma fruits naturally set parthenocarpically whereas Clementine mandarins show very low ability to set fruit in the absence of cross-pollination. The endogenous levels of gibberellins (GAs) and free and conjugated indole-acetic acid (IAA) and abscisic acid (ABA) throughout early stages of fruit development were investigated in seedless cultivars of both species. Analyses performed by full-scan combined gas chromatography-mass spectrometry (GC-MS) of extracts from ovaries at anthesis demonstrated the presence of GA₁₉, GA₂₀, GA₂₉, GA₁, GA₈, GA₃ and iso-GA₃ in Satsuma mandarin, whereas only GA₂₉, GA₃ and trace levels of GA₈ were detected in Clementine. At this developmental stage GA-like substances, as estimated by bioassay, reached their highest levels in Satsuma, while Clementine mandarins contained relatively lower levels. In both species the highest levels of free IAA were found at petal-fall stage at which time free ABA levels also peaked. Developing fruits of Clementine had higher amounts of both free IAA and ABA. In Satsuma, levels of conjugated IAA remained low throughout reproductive development whereas in Clementine they increased as the free form declined. In contrast, conjugated ABA was at low levels in Clementine but reached higher concentrations in Satsuma. These results suggest that in these mandarins the potential for setting parthenocarpic fruits is mainly influenced by the hormonal status of the fruit during the later stages of cell division and early stages of cell enlargement. Thus, the condition of low ability to set parthenocarpic fruits appears to be associated with lower levels of active GAs, lower capability to catabolize ABA to conjugated ABA and higher ability to conjugate IAA during this period.     

A ten-year study of population dynamics of citrus pests in the pesticide-reduced orchard

To determine the effectiveness of a pest management system that uses pesticides at a low level, we censused populations of seven insect pests and three plant diseases from 1980 to 1989 in a citrus orchard which was managed with a pesticide-reduced and pesticide-free protocols, in Wakayama Prefecture, Japan. In the orchard, we controlled the pests solely by spraying petroleum oil once a year from 1980 to 1985, by not spraying insecticide in 1986, and spraying only germicide in 1987. The arrowhead scale population remained much lower level at which citrus trees begin to wither when petroleum oil was sprayed, whereas it rapidly increased and began to wither trees during the years without spraying petroleum oil. The Indian wax scale population rapidly increased only during a 2-year period and then declined. Although the infection level of the sooty mold was unusually high on the trees where the density of the scale was high, this pathogen did little damage to citrus trees. The population of the red wax scale gradually increased throughout the census period but did not reach the level at which citrus trees begin to be damaged. Other pests also caused negligible damages to citrus trees. The results indicated that the pesticide-reduced pest management system which was proposed by Inoue and Ohgushi (1976, 1977) is valid for the prevention of the outbreak of the arrowhead scale, and that is has the sufficient control efficiency for the other pests in citrus orchards.     

Inhibition of the Acid Lipase Activity by Apolipoprotein A-I in the Presence of Lysosomal Proteases

It was found that the inhibition of the lysosomal acid lipase activity by rat apolipoprotein A-I (apo A-I) was increased with the degradation of apo A-I by the lysosomal proteases. We demonstrated that apo A-I could effectively inhibit the acid lipase activity even in the presence of the lysosomal proteases using the hepatic lysosomal fraction.     

温州蜜柑果实发育期间果糖激酶与糖积累的关系

研究了温州蜜柑果实发育进程中糖含量变化与果糖激酶活性变化的关系及增施氮肥对果实果糖激酶活性和基因表达的影响.结果表明,随着果实的发育,可食组织果糖激酶活性逐渐降低,糖含量不断增加,果皮中蔗糖和葡萄糖含量在成熟期略有下降,果糖激酶活性略有升高.果实膨大期后增施氮肥的果实在成熟期可食组织及果皮中蔗糖和果糖所占比例均有所下降,葡萄糖比例升高,以单位蛋白质表示的果糖激酶活性也明显高于对照果实.Northern分析表明,增施氮肥能促进发育后期果实可食组织中Cufrkl基因的表达,但对Cufrk2的表达无明显作用.     

Interaction between hyaluronate and the cell surface of Pseudomonas aeruginosa

Abstract Treatment of Pseudomonas aeruginosa cells with the non-metabolizable polysaccharide hyaluronate led to a strong increase in extracellular lipase activity. Alteration of the cell surface either by treatment with the chelator EDTA or by selecting for phage-resistant mutants significantly altered the bacterial response to hyaluronate. Binding of ¹⁴C-labeled hyaluronate to the bacteria was shown to depend on polysaccharide concentration and on cell number. Cell-free exolipase interacted with chemically cross-linked hyaluronate. The results suggested an interaction between hyaluronate and the cell surface of P. aeruginosa as a prerequisite for the polysaccharide to be effective.     

Interaction between Autonomous and Microtubule Guidance Systems Controls Cellulose Synthase Trajectories

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Interaction between cystatin-derived peptides and papain

The interaction between papain and synthetic peptides which tentatively mimic cystatin surfaces was investigated both enzymatically and structurally. Measurements of dissociation equilibrium constants for the interaction of papain with these peptides modified by successive deletions or substitutions demonstrated that the QVVAG segment, which is highly conserved throughout members of the cystatin superfamily, is essential for the interaction. The glycylcontaining (N-terminal) fragments and PW-containing (C-terminal) fragments were found to be of lesser importance, since each could be deleted without significantly modifying the interaction. These fragments improved the stability of the interacting QVVAG region, which appeared to be substrate-like in all peptides tested, as it was cleaved at the A-G bond upon peptide-papain interaction. Replacement of the A residue at the scissile bond of the QVVAG by a blocked cysteinyl residue reduced the rate of cleavage of the susceptible bond and therefore shifted the resulting peptide from a substrate to an inhibitor. Derivatization of this substituted peptide at its N- and C-terminal ends by fluoresceinyl groups resulted in a dramatic decrease in theK _i to 0.5 µM. This improvement in the inhibitory properties of the substituted and derivatized peptides was correlated with structural changes as analyzed by molecular dynamic calculations. The results were compared to those proposed for the mechanism of inhibition by natural inhibitors of the cystatin superfamily.     

Interaction between the basic inhibitor of bovine pancreas and chymotrypsin and trypsin. Selective anthraniloylation of the maleylated inhibitor

The interaction between the maleylated basic pancreatic inhibitor, anthraniloylated on its lysine-15 residue, and chymotrypsin is studied by fluorescence intensity, fluorescence polarization, circular dichroism, circular polarization of fluorescence and sedimentation. These measurements show that the interaction takes place through the entrance of the anthraniloyl group into an asymmetric environment in which it is rigidly held. The dissociation constant of the complex is 2.5 × 10^?8m. The interaction between the modified inhibitor and trypsin takes place through a site which is not the anthraniloylated lysine-15 side-chain, yet not far from it.