Oligomeric proanthocyanidins from Rumex acetosa L. inhibit the attachment of herpes simplex virus type-1

The polyphenole-enriched acetone-water extract R2 from the aerial parts of Rumex acetosa L. containing high amounts of oligomeric and polymeric proanthocyanidins and flavonoids was tested for antiviral activity. R2 exhibited strong antiviral activity against herpes simplex virus type-1 (HSV-1) while the replication of adenovirus 3 was not affected. By plaque reduction test and MTT assay on Vero cells, the HSV-1-specific inhibitory concentration (IC(50)) and cytotoxic concentration (CC(50)) were determined. R2 exibited an IC(50) of 0.8 μg/mL and a selectivity index (SI) (ratio of IC(50) to CC(50)) of approximately 100 when added to the virus inoculum for 1h at 37°C prior to infection. The antiviral activity was due to the presence of flavan-3-ols and oligomeric proanthocyanidins in the extract. Structure-activity analyses indicated that flavan-3-ols and proanthocyanidins with galloylation at position O-3 are highly potent compounds (SI>40), while ungalloylated compounds did not exhibit antiviral effects (SI<1). R2 and a major proanthocyanidin from R2, epicatechin-3-O-gallate-(4β→8)-epicatechin-3-O-gallate abolished virus entry into the host cell by blocking attachment to the cell surface. When added after attachment at a concentration of ≥ 12.5 μg/mL, R2 inhibited also penetration of HSV-1 into the host cell. R2 and epicatechin-3-O-gallate-(4β→8)-epicatechin-3-O-gallate were shown to directly interact with viral particles leading to the oligomerisation of envelope proteins as demonstrated for the essential viral glycoprotein gD. Using raft cultures with three-dimensional organotypic human skin equivalents it was shown that treatment of cultures with R2 after infection with HSV-1 resulted in a reduced viral spread.     

A-type procyanidin derivatives with antioxidant and much enhanced α-glucosidase inhibitor activities

Procyanidins are natural compounds with good biological activity. However, due to a large number of phenolic hydroxyl groups in the structure, they have high polarity and low bioavailability. The preparation of A-type proanthocyanidin derivatives is an effective way to change their polarity and biological activity. In this paper, a series of A-type procyanidin derivatives were designed and synthesized by two practical and safe methods, and two new dimeric A-type procyanidin derivatives, procyanidin A1-acetone conjugate (6) and procyanidin A2-cystein conjugate (9) were obtained and reported for the first time. Their structures were characterized and confirmed by ¹H NMR, ¹³C NMR, HMBC, ¹H-¹H COSY and MS. All the compounds showed strong DPPH scavenging activities. Compound 6 showed inhibitory effects on α-glucosidase with an IC₅₀ value of 8.7 μg/mL, while its parental compound, procyanidin A1, had no inhibitory effects. Degradation of procyanidins from peanut skin by L-cystein was studied. The results showed that the main structural unit of procyanidins in peanut skin was A-type proanthocyanidins dimer.     

Anti-inflammatory and anti-melanogenic steroidal saponin glycosides from Fenugreek (Trigonella foenum-graecum L.) seeds

Fenugreek seed ( Trigonella foenum-graecum L.) is used as an herbal medicine for treating metabolic and nutritive dysfunctions. To determine if this plant has other beneficial effects, we tested the inhibitory activities of a methanol (MeOH) extract of fenugreek seed on the production of inflammatory cytokines and melanin synthesis in cultured cell lines in vitro. The MeOH extract inhibited the production of phorbol-12-myristate-13-acetate-induced inflammatory cytokines such as tumor necrosis factor (TNF)-α in cultured THP-1 cells, and also restrained the intracellular synthesis of melanin in murine melanoma B16F1 cells. We isolated three active constituents from fenugreek seed extracts. These were identified as the steroidal saponins 26- O-β-D-glucopyranosyl-(25 R)-furost-5(6)-en-3 β,22 β,26-triol-3- O-α-L-rhamno-pyranosyl-(1' → 2')-O-[β-D-glucopyranosyl-(1' → 6')- O]-β-D-glucopyranoside 1, minutoside B 2, and pseudoprotodioscin 3. Compounds 1 and 2 strongly suppressed the production of inflammatory cytokines, whereas 3 showed a weaker suppressing effect. Melanogenesis in B16F1 cells was significantly suppressed by 1 and 3, and weakly suppressed by 2. All three compounds showed moderate cytotoxicities. These results indicate that fenugreek extract and its active constituents could protect against skin damage.     

(1→3)-β-d-Glucan stimulates nitric oxide generation and cytokine mRNA expression in macrophages

Beta-glucans are known for their potent ability to induce nonspecific inflammatory reactions and are believed to play a role in bioaerosol-induced respiratory symptoms seen in both occupational and residential environments. Here, the ability of a (1→3)-β-d-glucan (Curdlan) to stimulate nitric oxide generation and cytokine mRNA expression in rat alveolar macrophages (AMs) and the murine monocyte/macrophage cell line, RAW 264.7 was investigated. Exposure to (1→3)-β-d-glucan (20, 100 and 500 μg/ml) induced a dose-dependent increase in the expression of inducible nitric oxide synthase mRNA and a release of nitric oxide into the culture medium in both rat AMs and RAW 264.7 cells. The mRNA expression of a number of other inflammatory mediators such as interleukin-1β, interleukin-6, tumor necrosis factor-α and cyclooxygenase-2 was also increased by the exposure to β-glucan. The capability of (1→3)-β-d-glucan (500 μg/ml) to induce mRNA synthesis of these various mediators were comparable to that of endotoxin (1 μg/ml). These results imply that (1→3)-β-d-glucan stimulates the generation of nitric oxide, cytokines and prostaglandins in macrophages and suggest the possibility that this may contribute to bioaerosol-induced respiratory symptoms seen in exposed individuals.     

异甘草酸镁对人表皮角质形成细胞炎症因子的产生和外周血白细胞趋化功能的影响

目的：研究异甘草酸镁对角质形成细胞产生促炎症细胞因子、黏附分子和对白细胞趋化功能的影响,以获得异甘草酸镁治疗炎症性皮肤病的药理学基础.方法：采用2 000 U/mL rhIFN-γ诱导人表皮角质形成细胞Ha-CaT细胞,用ELISA法检测不同浓度的异甘草酸镁对HaCaT细胞IL-8、IL-6和ICAM-1产生的影响;通过琼脂糖平板法在显微镜下观察药物对10 nmol/L甲酰甲二磺酰-亮氨酰-苯丙氨酸（fMLP）诱引的人白细胞移动距离的影响.结果：异甘草酸镁能剂量依赖性地抑制rhIFN-γ刺激引起的IL-8、IL-6和ICAM-1的产生,其IC50值分别为1.21、7.23和4.17μg/mL;对fMLP诱引的人外周血白细胞趋化功能有明显影响,其抑制作用呈剂量依赖性,IC50值为8.58 μg/mL.结论：异甘草酸镁可通过抑制人表皮角质形成细胞IL-8、IL-6、ICAM-1产生和白细胞趋化达到治疗炎症相关性皮肤病的作用.     

Aqueous extract of peanut skin and its main constituent procyanidin A1 suppress serum IgE and IgG1 levels in mice-immunized with ovalbumin

In this study, we investigated the effects of an aqueous extract of peanut (Arachis hypogaea L.) seed skin (PSE) and its main constituent procyanidin A1 (PA) on the allergic response to allergen ovalbumin (OVA) in a mouse model. Mice immunized interaperitoneally with OVA dramatically increased anti-OVA IgE and total IgG1 levels in serum compared with non-treated control mice. Oral injection of PSE at doses ranging from 10 to 100 mg/kg/d (for 21 consecutive days) decreased anti-OVA IgE and IgG1 levels 21 d after OVA-immunization. OVA-induced increments in spleen weight and peripheral white blood cell count were also suppressed by this PSE administration. Polyphenol-enriched fractions from apple (30 mg/kg) and grape seed (30 mg/kg) also decreased anti-OVA IgE level but did not affect total IgG1 levels. Oral injection of PA (1 to 10 mg/kg/d) purified from PSE resulted in a suppression of IgE and total IgG1 levels in serum. An increment of serum interleukin-4 level in mice that were immunized with OVA was reduced by all tested samples, whereas PSE and PA were the only compounds that could reverse the reduced interferon-gamma level by OVA. These findings suggest that intake of PSE or its main active constituent PA may prevent an allergic reaction by inhibiting immunoglobulin synthesis, and the mechanism of this action of PSE and PA is in part due to their regulation of T helper cytokine production.     

多波长高效液相色谱法同时测定夜宁合剂中4种有效成分的含量

目的建立多波长HPLC梯度洗脱法测定夜宁合剂中(-)-丁香树脂酚-4-O-β-D-呋喃芹糖基-(1→2)-β-D-吡喃葡萄糖苷、女贞苷、特女贞苷和槲皮苷的含量。方法采用C18色谱柱(250 mm×4.6mm,5μm);流速:1.2 mL·min-1;流动相A为乙腈-甲醇(1:2),B为0.1%磷酸溶液,梯度洗脱;检测波长:(-)-丁香树脂酚-4-O-β-D-呋喃芹糖基-(1→2)-β-D-吡喃葡萄糖苷为204 nm,女贞苷和特女贞苷为224 nm,槲皮苷为360 nm,柱温为30℃。结果 (-)-丁香树脂酚-4-O-β-D-呋喃芹糖基-(1→2)-β-D-吡喃葡萄糖苷、女贞苷、特女贞苷和槲皮苷质量浓度分别在5.75¹15.00μg·mL-1(r=0.999 7)、6.45115.00μg·mL-1(r=0.999 7)、6.45¹29.00μg·mL-1(r=0.9995)、5.30129.00μg·mL-1(r=0.9995)、5.30¹06.00μg·mL-1(r=0.999 9)、8.45106.00μg·mL-1(r=0.999 9)、8.45¹69.00μg·mL-1(r=0.999 8)与峰面积线性关系良好;(-)-丁香树脂酚-4-O-β-D-呋喃芹糖基-(1→2)-β-D-吡喃葡萄糖苷、女贞苷、特女贞苷和槲皮苷的平均加样回收率分别为97.7%、96.8%、98.8%、98.2%,RSD(n=6)分别为1.5%、0.60%、1.1%、1.1%。结论该方法快速、灵敏、准确,可作为夜宁合剂的含量控制方法。     

Testosterone replacement therapy promotes angiogenesis after acute myocardial infarction by enhancing expression of cytokines HIF-1a, SDF-1a and VEGF

Soluble amyloid β-protein (Aβ) oligomers are primary mediators of synaptic dysfunction associated with the progression of Alzheimer's disease. Such Aβ oligomers exist dependent on their rates of aggregation and metabolism. Use of selective somatostatin receptor-subtype agonists have been identified as a potential means to mitigate Aβ accumulation in the brain, via regulation of the enzyme neprilysin. Herein, we first evaluated the impact of the somatostatin receptor subtype-4 agonist 1-[3-[N-(5-Bromopyridin-2-yl)-N-(3,4-dichlorobenzyl)amino]propyl]-3-[3-(1H-imidazol-4-yl)propyl]thiourea (NNC 26-9100) on learning and memory in 12-month SAMP8 mice (i.c.v. injection). NNC 26-9100 (0.2 μg-dose) was shown to enhance both learning (T-maze) and memory (object recognition) compared to vehicle controls. Cortical and hippocampal tissues were evaluated subsequent to NNC 26-9100 (0.2 μg) or vehicle administration for changes in neprilysin activity, along with protein expression of amyloid-precursor protein (APP), neprilysin, and Aβ???? oligomers within respective cellular fractions (extracellular, intracellular and membrane). NNC 26-9100 increased neprilysin activity in cortical tissue, with an associated protein expression increase in the extracellular fraction and decreased in the intracellular fraction. A decrease in intracellular APP expression was found with treatment in both cortical and hippocampal tissues. NNC 26-9100 also significantly decreased expression of Aβ???? trimers within both the extracellular and intracellular cortical fractions. No expression changes were found in membrane fractions for any protein. These finding suggest the potential use of selective SSTR4 agonists to mitigate toxic oligomeric forms of Aβ???? in critical regions of the brain identified with learning and memory decline.     

头孢拉定原料及制剂的聚合物杂质分析#br#

目的 建立头孢拉定原料及制剂中聚合物杂质的分析方法。方法 采用碱降解法制备头孢拉定强制降解溶液;采用高效凝胶色谱法(TSK G2000 SWxl)和柱切换-LC/MS法对头孢拉定强制降解溶液中的聚合物杂质进行分离和结构鉴定;采用Agilent ZORBAX SB-C18型色谱柱,以磷酸盐缓冲液-甲醇为流动相,进行梯度洗脱,建立头孢拉定聚合物的RP-HPLC分析方法,采用二维液相色谱法和柱切换-LC/MSn法对该方法的专属性进行分析;进行方法学验证。结果 在头孢拉定强制降解物中鉴定出头孢拉定二聚体、三聚体、四聚体;高效凝胶色谱法分离头孢拉定聚合物杂质时,易受到小分子杂质的干扰,同时分离的聚合物杂质色谱峰拖尾严重,存在肩峰,定量准确性差;RP-HPLC法分析头孢拉定聚合物杂质时,在20~22min范围内检出头孢拉定二聚体、三聚体、四聚体;方法定量限为500μg,最低检测限为150μg。结论 高效凝胶色谱法不能对头孢拉定中的聚合物杂质进行有效质控,建立的反相色谱法分析头孢拉定聚合物杂质时专属性良好、灵敏度高、方法耐用性好,可用于头孢拉定原料及制剂的聚合物杂质质控;头孢拉定强制降解溶液可作为头孢拉定聚合物分析的系统适用性溶液。     

Isoforskolin pretreatment attenuates lipopolysaccharide-induced acute lung injury in animal models

Isoforskolin was isolated from Coleus forskohlii native to Yunnan in China. We hypothesize that isoforskolin pretreatment attenuates acute lung injury induced by lipopolysaccharide (endotoxin). Three acute lung injury models were used: situ perfused rat lung, rat and mouse models of endotoxic shock. Additionally, lipopolysaccharide stimulated proinflammatory cytokine production was evaluated in human mononuclear leukocyte. In situ perfused rat lungs, pre-perfusion with isoforskolin (100, and 200 μM) and dexamethasone (65 μM, positive control) inhibited lipopolysaccharide (10 mg/L) induced increases in lung neutrophil adhesion rate, myeloperoxidase activity, lung weight Wet/Dry ratio, permeability-surface area product value, and tumor necrosis factor (TNF)-α levels. In rats, pretreatments with isoforskolin (5, 10, and 20 mg/kg, i.p.) and dexamethasone (5mg/kg, i.p.) markedly reduced lipopolysaccharide (6 mg/kg i.v.) induced increases of karyocyte, neutrophil counts and protein content in bronchoalveolar lavage fluid, and plasma myeloperoxidase activity. Lung histopathology showed that morphologic changes induced by lipopolysaccharide were less pronounced in the isoforskolin and dexamethasone pretreated rats. In mice, 5 mg/kg isoforskolin and dexamethasone caused 100% and 80% survival, respectively, after administration of lipopolysaccharide (62.5mg/kg, i.v., 40% survival if untreated). In human mononuclear leukocyte, isoforskolin (50, 100, and 200 μM) and dexamethasone (10 μM) pre-incubation lowered lipopolysaccharide (2 μg/mL) induced secretion of the cytokine TNF-α, and interleukins (IL)-1β, IL-6, and IL-8. In conclusion, pretreatment with isoforskolin attenuates lipopolysaccharide-induced acute lung injury in several models, and it is involved in down-regulation of inflammatory responses and proinflammatory cytokines TNF-α, IL-1β, IL-6, and IL-8.     

酮洛芬脂质体凝胶在大鼠体内的药物动力学及皮肤组织分布研究

目的：研究酮洛芬（KPF）脂质体凝胶在大鼠体内的药物动力学及其皮肤局部组织分布。方法：选择SD大鼠为动物模型,与KPF口服给药相对照,研究KPF脂质体凝胶局部用药后在血浆和皮肤中的体内药动学行为。结果：KPF脂质体凝胶大鼠局部给药后的血药浓度Cmax、AUC0→24分别为（1．255±0．54）μg·ml^-1和（59．388±3．76）μg·ml^-1·h,局部皮肤组织的Cmax、AUC0→24分别为（28．019±1．98）μg·ml^-1和（602．21±2．84）μg·ml^-1·h。与口服KPF相比,KPF脂质体凝胶降低了血液中的分布,而具有明显的皮肤蓄积能力（P〈0．05）。结论：KPF脂质体凝胶局部给药在获得较高的皮肤组织浓度的同时,避免了较高的血药浓度的不良反应,疗效优于口服．     

Effects of grape seed proanthocyanidin on 5-hydroxytryptamine(3) receptors in NCB-20 neuroblastoma cells

Proanthocyanidin is a phenolic compound present in plants, that has antioxidant, antinociceptive, anti-emetic, and neuroprotective properties. We investigated the actions of proanthocyanidin from grape seeds on 5-hydroxytryptamine (5-HT)(3) receptors in NCB-20 neuroblastoma cells using a whole-cell voltage clamp technique. Co-treatment of proanthocyanidin (0.3-100 μg/ml) and 3 μM 5-HT (near EC(50)) produced a slight inhibition of 5-HT-induced inward peak current (I(5-HT)) in NCB-20 cells, but pretreatment with proanthocyanidin for 30 s before application of 5-HT induced a much larger inhibition of I(5-HT) in an irreversible, concentration- and time-dependent manner (IC(50)=6.5±0.4 μg/ml, Hill coefficient=2.5±0.1). Proanthocyanidin also produced a concentration-dependent inhibition of currents induced by 30 μM 5-HT, near-maximal concentration (IC(50)=22.1±0.4 μg/ml, Hill coefficient=2.4±0.1). High concentrations (≧30 μg/ml) of proanthocyanidin caused a concentration-dependent inhibition of the activation and desensitization of currents induced by 30 μM 5-HT. Further studies showed that pretreatment of 20 μg/ml proanthocyanidin caused not only a rightward shift of the dose-response curve for 5-HT (EC(50) shift from 2.7±0.4 to 6.2±0.5 μM), but also a decreased E(max) (inhibition by 37.5±1.3%). The proanthocyanidin-induced inhibition of 5-HT(3) receptors did not show a significant difference within the testing holding potential ranges (-50-+30 mV). These results suggest that proanthocyanidin inhibits 5-HT(3) receptor function in NCB-20 cells in a noncompetitive mode, and that this inhibitory effect of proanthocyanidin probably contributes to the pharmacological actions of proanthocyanidin.     

Determination of the oestrogenic (uterotrophic) activity of extracts of 'general purpose polystyrene (GPPS)' using immature female rats

In Japan there is growing concern about the possible adverse effects of consumption of food from styrene containers (mainly those made from polystyrene paper) due to the alleged oestrogenic activity of styrene oligomers (dimers and trimers), which may migrate into the food. To examine the possible oestrogenic activity of styrene dimers and trimers, extracts were made from 'general purpose polystyrene (GPPS)' and administered orally to immature female rats over a 4 day period. Increase of uterus weight (wet and blotted) was used for assessment of possible oestrogenic activity. To establish the sensitivity of the test method, immature rats were treated with diethylstilboestrol (DES), a well-known oestrogenic compound. It was found that treatment of rats with levels of up to 60 microg of styrene dimers and 930 microg of styrene trimers per kilogram body weight per day did not give any statistically significant increase of the uterus weight (wet or blotted), whereas DES caused statistically significant, dose-related increases in uterus weight at levels as low as 0.89 microg kg(-1) body weight day(-1). It was concluded that, compared with the estimated maximum human daily intake of styrene trimers of 1 microg kg(-1) body weight day(-1) from polystyrene food containers, the risk of adverse human health effects with respect to oestrogenicity may be considered negligible.     

HPLC波长切换法同时测定理气散结颗粒中4个成分的含量

目的建立同时测定理气散结颗粒中氧化芍药苷、芍药内酯苷、芍药苷和α-香附酮的HPLC波长切换联合梯度洗脱法。方法采用Venusil MP C18色谱柱(4.6 mm×250 mm,5μm),以乙腈(A)-0.1%磷酸溶液(B)为流动相,梯度洗脱(0~15 min,12.0%A;15~31 min,12.0%A→26.0%A;31~39 min,26.0%A→40.0%A;39~45 min,40.0%A→12.0%A),流速0.9 mL/min,波长切换(0~31 min,在230 nm波长下检测氧化芍药苷、芍药内酯苷和芍药苷;31~45 min,在242 nm波长下检测α-香附酮),柱温30℃,进样量为20μL。结果氧化芍药苷、芍药内酯苷、芍药苷和α-香附酮4个成分的质量浓度分别在5.15~103.00μg/mL(r=0.999 6)、8.99~179.80μg/mL(r=0.999 5)、19.83~396.60μg/mL(r=0.999 9)、6.35~127.00μg/mL(r=0.999 3)呈良好线性关系;平均加样回收率及相应的RSD(n=6)分别为96.83%(0.93%)、97.85%(1.31%)、99.71%(0.80%)、98.77%(1.59%)。结论所建立的方法灵敏度高、快速、专属性好、准确度高,为理气散结颗粒的质量控制提供了依据。     

Multiwalled carbon nanotubes induce a fibrogenic response by stimulating reactive oxygen species production, activating NF-κB signaling, and promoting fibroblast-to-myofibroblast transformation

Carbon nanotubes (CNTs) are novel materials with unique electronic and mechanical properties. The extremely small size, fiberlike shape, large surface area, and unique surface chemistry render their distinctive chemical and physical characteristics and raise potential hazards to humans. Several reports have shown that pulmonary exposure to CNTs caused inflammation and lung fibrosis in rodents. The molecular mechanisms that govern CNT lung toxicity remain largely unaddressed. Here, we report that multiwalled carbon nanotubes (MWCNTs) have potent, dose-dependent toxicity on cultured human lung cells (BEAS-2B, A549, and WI38-VA13). Mechanistic analyses were carried out at subtoxic doses (≤20 μg/mL, ≤ 24 h). MWCNTs induced substantial ROS production and mitochondrial damage, implicating oxidative stress in cellular damage by MWCNT. MWCNTs activated the NF-κB signaling pathway in macrophages (RAW264.7) to increase the secretion of a panel of cytokines and chemokines (TNFα, IL-1β, IL-6, IL-10, and MCP1) that promote inflammation. Activation of NF-κB involved rapid degradation of IκBα, nuclear accumulation of NF-κBp65, binding of NF-κB to specific DNA-binding sequences, and transactivation of target gene promoters. Finally, MWCNTs induced the production of profibrogenic growth factors TGFβ1 and PDGF from macrophages that function as paracrine signals to promote the transformation of lung fibroblasts (WI38-VA13) into myofibroblasts, a key step in the development of fibrosis. Our results revealed that MWCNTs elicit multiple and intertwining signaling events involving oxidative damage, inflammatory cytokine production, and myofibroblast transformation, which potentially underlie the toxicity and fibrosis in human lungs by MWCNTs.     

HPLC波长切换法同时测定藤药中5种成分的含量

目的建立同时测定藤药中5种成分含量的HPLC方法。方法采用Phenomene C18色谱柱(4.6 mm×250 mm,5μm)。流动相采用甲醇(A)-水(B)梯度洗脱:0~15 min(45.0%A),15~27 min(45.0%→56.0%A),27~42 min(56.0%→70.0%A),42~50 min(70.0%→45.0%A)。流速0.8 m L/min;柱温25℃;检测波长:0~15 min(248 nm,α-玉柏碱),15~50 min(254 nm,升麻素苷、升麻素、5-O-甲基维斯阿米醇苷和亥茅酚苷)。结果测得α-玉柏碱、升麻素苷、升麻素、5-O-甲基维斯阿米醇苷和亥茅酚苷的平均加样回收率及对应的RSD分别为98.63%(1.60%)、97.82%(1.45%)、99.18%(1.48%)、97.18%(1.29%)和98.19%(1.80%),且在5.15~103.00、8.13~162.60、4.74~94.80、4.09~81.80、4.35~87.00μg/m L内,线性关系良好(r≥0.999 1)。结论采用HPLC波长切换法能有效分析藤药中的5个成分,方法准确可靠,重复性良好,结果稳定,可以作为藤药产品质量控制的方法。     

Temperature regulates melanin synthesis in melanocytes

Temperature change is one of the major environmental factors that influence the human skin. However, the relationship between temperature and melanogenesis has received little attention. In the present study, we investigated the effects of temperature change on melanogenesis in a mouse melanocyte cell line (Mel-Ab), and primary cultured human melanocytes. We found that Mel-Ab cells cultured at low temperatures (31 and 34 degrees C) produce less melanin than cells at 37 degrees C. These results were confirmed by experiments upon human melanocytes, demonstrating that the hypopigmenting effect of low temperatures is not cell type dependent. The observed melanin production was found to be accompanied by tyrosinase activity at each temperature, indicating that tyrosinase activity is regulated by temperature. We further examined whether the incubation period at low temperatures plays an important role in the regulation of melanogenesis. Short exposures to 27 degrees C for 1 h or 3 h did not affect tyrosinase activity or melanin synthesis, whereas long exposures to 31 degrees C for 2 days or 6 days significantly reduced tyrosinase activity and melanin synthesis in a duration-dependent manner. Our results suggest that exposure to low temperature and the duration of this exposure are important regulators of melanogenesis.     

4味中药乙醇提取物祛黄褐斑作用的比较研究

目的比较白附子、银花、白芨、六月雪4味中药乙醇提取物祛黄褐斑作用的差异,选出作用最强的药材。方法以MTT法测定小鼠B16黑素瘤细胞增殖抑制率、酪氨酸酶活性、黑素含量变化;采用多重比较法进行统计分析。结果银花的浓度为62.5μg.mL-1,白芨、白附子各浓度(500、250、125、62.5μg.mL-1)均对B16鼠黑素瘤细胞增殖抑制作用较强。白附子的浓度为250μg.mL-1或银花的浓度为62.5μg.mL-1对小鼠B16黑素瘤细胞酪氨酸酶活性抑制作用最佳。白附子的浓度为125μg.mL-1对黑素合成抑制作用最好。结论白芨与白附子可作为祛黄褐斑有效成分的候选药材。     

Silymarin inhibits melanin synthesis in melanocyte cells

Objectives The aim was to search for inhibitors of melanogenesis from natural resources. Methods The inhibitory effect of silymarin on melanogenesis in a spontaneously immortalized mouse melanocyte cell line, Mel‐Ab, was studied. Key findings Silymarin significantly prevented melanin production in a dose‐dependent manner with an IC50 value (concentration producing 50% maximal inhibition) of 28.2 μg/ml, without effects on cell viability. Also, silymarin inhibited l ‐DOPA oxidation activity of tyrosinase, the rate‐limiting melanogenic enzyme, in cell based‐systems but it did not directly affect cell‐free tyrosinase activity. Furthermore, Western blot analysis indicated that silymarin decreased the expression of tyrosinase protein. Conclusions This study suggests that the depigmenting effect of silymarin might be attributable to inhibition of tyrosinase expression and that silymarin may be useful as a natural skin‐lightening agent.