牛角膜混浊和渗透性试验方法与家兔法评价染发类化妆品眼刺激性的比较研究

目的 探讨在染发类化妆品备案审查中,用牛角膜混浊和渗透性试验(BCOP)代替家兔法(Draize 试验)进行眼刺激性评价的可能性。方法 分别用BCOP与Draize 试验对14种市售染发剂进行眼刺激性评价,并对评价结果进行比较。结果 对14种染发剂眼刺激性检测的结果表明,Draize 试验所得的刺激反应等级为微刺激性―轻刺激性,按现行评审标准,通过率为100%(14/14);BCOP所得的刺激反应等级为轻微刺激性―严重刺激性,与Draize法分类一致性达到78.6%(11/14),以体外刺激评分值(IVIS)≤25为限值,通过率为78.6%(11/14);以IVIS≤55为限值,通过率为85.7%(12/14)。结论 用BCOP代替Draize 试验对染发类化妆品进行眼刺激性评价可行。     

化妆品的眼刺激性实验研究

化妆品因其广泛的使用范围及日益巨大的使用数量 ,使化妆品使用的安全性问题显得越来越重要。化妆品的眼刺激性和腐蚀性是控制化妆品卫生质量的一项重要的毒理学指标。一年来 ,我们根据《化妆品卫生规范》(1999)对 32 6种样品进行了急性眼刺激性试验 ,现将结果报道如下。1　材料与方法1 1　实验材料　送检、受理申报的国产或进口的各类用于眼部及易接触眼的化妆品 ,包括眼部护肤类、洗面奶类、洗发沐浴类、护发美发类等普通化妆品 173种 ,染发剂等特殊用途化妆品共 15 3种 ,所有样品均不稀释 (染发剂等需二剂及以上混合后方可使用的产品 ,按…     

BCOP法评价滴眼液的角膜渗透性

目的 采用牛角膜混浊和渗透性试验（bovine corneal opacity and permeability,BCOP）方法对滴眼液的角膜渗透性进行评价。方法 利用BCOP法,以无酚红MEM培养液为介质,前室加滴眼液样品,于不同时间点后室取样,用紫外-可见分光光度法测定其苄达赖氨酸的含量,比较不同滴眼液样品的角膜渗透性。结果 不同厂家样品表现出不同的角膜渗透性,包括角膜累积渗透量、透过速率、表观渗透系数和稳态渗透速率均不同。结论 可用BCOP法进行眼用制剂的角膜渗透性研究。     

应用BioOcullarTM重建人角膜上皮模型评估化学物和化妆品产品眼刺激性

目的采用BioOcullar^TM重建人角膜上皮模型检测化学物和化妆品产品的眼刺激性,以评价基于该模型的测试方法对化学物和化妆品眼刺激性评估能力。方法检测化学物时,选用58种化学物,暴露于角膜上皮模型后,用噻唑蓝(MTT)法测定角膜组织活性,计算组织相对活性值,当组织相对活性≤60%时,判定受试物为刺激性阳性,当组织相对活性>60%时,判定为刺激性阴性。检测化妆品产品时,选用61种不同类型化妆品,采用时间毒性效应二步法,根据测试结果划定刺激性分级标准。分析测试方法对化学物和化妆品眼刺激性检测的灵敏度、特异度和符合率。结果对58种化学物的眼刺激性检测,其中25种化学物3轮重复测试刺激性分类均为一致,且与欧洲化学品管理局分类、标签和包装数据库(Classification,Labelling&Packaging,CLP)或全球化学物统一分类和标签系统(Globally Harmonized System of Classification and Labelling of Chemicals,GHS)分类一致;扩大对33种化学物检测,有32种结果与CLP或GHS分类结果一致,对58种化学物眼刺激性检测的灵敏度为100%,特异度为95.00%,符合率为98.28%。对61种化妆品1 h筛检结果,有26份化妆品划分为无刺激性或微刺激性,符合率为100%;对余下的35份疑似刺激性化妆品检测,参照动物试验结果划定刺激性分级标准为:ET50>36.5 min为无刺激或微刺激性,5 min     

鸡胚绒毛膜尿囊膜血管试验法用于评价化妆品眼刺激性的可行性研究

目的 研究鸡胚绒毛膜尿囊膜血管试验法（chorioallantoic membrane vascular assay,CAMVA）评价化妆品眼刺激性的可行性,并用改良后的方法对14种市售化妆品进行眼刺激性的评价。方法 通过控制样品的浓度、与鸡胚绒毛膜尿囊膜的接触时间,以及对评价体系的改良,尝试建立一套标准的CAMVA法评价化妆品眼刺激性的方法。结果 护肤类产品无刺激性,用后冲洗类、染发类产品的浓度、与鸡胚绒毛膜尿囊膜的接触时间均与刺激性大小成正比。在3%的浓度下,用后冲洗类产品的最佳接触时间为20 min,体内外试验结果一致性为90%（9/10）;染发类产品为5 min,体内外试验结果一致性为100%（2/2）。结合改良后的评价体系可做到刺激性分级,体内外试验结果一致性为用后冲洗类产品80%（8/10）,染发类产品50%（1/2）。结论 用CAMVA法可以对化妆品的眼刺激性进行评价,但实验条件和判定标准还需进一步优化。     

BioOcularTM角膜上皮模型EIT法评价眼用制剂包装材料的眼刺激性研究

目的：用BioOcularTM 角膜上皮模型EIT法评价眼用制剂包装材料的眼刺激性。方法：采用BioOcularTM 角膜上皮模型EIT法对6批眼用制剂包装材料的眼刺激性进行检测,并与Draize试验所得的结果相比较。结果：Draize法和BioOcularTM 角膜上皮模型EIT法所得眼刺激性结果均为无刺激性,与Draize法比较,BioOcularTM 角膜上皮模型EIT法与体内试验的分类一致性为100%(6/6)。 结论：三维重组角膜上皮模型 EIT法有潜力作为眼用制剂包装材料眼刺激性试验的替代方法。     

重组人皮肤模型在化妆品皮肤刺激性评价中的应用研究

目的 探讨重组人皮肤模型应用于化妆品体外皮肤刺激性评价的可行性.方法 以重组人皮肤模型(EpiSkinTM)为受试模型,在对10个已知刺激性分类的标准化学品进行方法验证的基础上,再根据化妆品的使用特性对23个产品进行皮肤刺激性分类评价,其中6个基础型化妆品和10个美容型化妆品暴露18 h、7个清洁型化妆品分别暴露18、...     

鸡胚绒毛膜尿囊膜血管试验用于评价化妆品配方眼刺激性质量标准的建立与评价

目的 建立鸡胚绒毛膜尿囊膜血管试验用于评价化妆品配方眼刺激性的标准。方法 通过对鸡胚绒毛膜尿囊膜血管试验原有判定方法的改进,建立针对化妆品配方眼刺激性评价的质量标准,以此对10种不同类型共20批次的化妆品配方进行眼刺激性评价,并将结果与家兔法相比较,验证质量标准的可行性。结果 20批次的化妆品配方鸡胚绒毛膜尿囊膜血管试验结果与家兔法相比一致率为85%（17/20）。结论 所建立的标准可用于化妆品配方眼刺激性评价。     

三维重组眼角膜上皮模型法与兔眼刺激法对洗发水眼刺激性的比较与评价

目的 本文同时采用三维人重组角膜模型BioOcular?和现行的兔眼刺激Daize法,对市面常见的不同配方洗发水的眼刺激性进行评价与比较。方法 参考体外眼刺激实验(OECD指导原则492),采用人重组眼角膜上皮模型和兔眼刺激法对8种不同品牌洗发水的眼刺激性进行评价研究。结果 研究表明,与传统兔眼实验比较,采用皮肤模型的眼刺激法对8种洗发水有无刺激性的区分结果与兔眼刺激法基本一致。其中两种方法相关系数r达到0.955,具有很好的相关性。结论 与兔眼刺激法相比,人重组眼角膜上皮模型刺激实验所需的周期短、试验步骤相对简便、检测结果相对更加客观,人源重组组织与人眼刺激更为接近,是一种值得推荐的眼刺激替代检测方法。     

Improvement of the Bovine Corneal Opacity and Permeability (BCOP) assay as an in vitro alternative to the Draize rabbit eye irritation test

Measurement of ocular irritancy is a necessary step in the safety evaluation of both industrial and consumer products. Assessment of the acute eye irritation potential is therefore part of the international regulatory requirements for testing of chemicals.The Bovine Corneal Opacity and Permeability (BCOP) assay is generally accepted as a valid in vitro alternative method to the Draize eye irritation test to detect corrosive and severe eye irritants (category 1), but has not proven sensitive enough to discriminate accurately moderate (category 2A/2B) to mild and non-irritating compounds. In the currently accepted BCOP assay, opacity is determined by the amount of light transmission through the cornea, and permeability is determined by the amount of sodium fluorescein dye that passes through all corneal cell layers. Both measurements are used to assign an In Vitro Irritancy Score (IVIS) for prediction of the in vivo ocular irritation potential of a test substance. Nowadays, opacity is measured by an OP-KIT opacitometer providing a center-weighted reading of light transmission by measuring changes in voltage when the transmission of white light passes through the cornea alters. As a consequence, this may underestimate opacity that develops as spots or heterogeneous opaque areas on the periphery of an isolated cornea.A prototype of a laser light-based opacitometer (PLLBO) allowing better measurement of opacities was developed by Van Goethem et al. (2010). This new device showed improved sensitivity to detect subtle changes in corneal transparency. Furthermore, the new opacitometer allowed the analysis of the complete corneal surface and was able to detect more efficiently opaque spots located along the sides of the excised corneas.A further improved prototype of the PLLBO was constructed in combination with a camera and a speckle noise reducer. Treatment conditions of the corneas in the cornea holders were optimized in order to mimic more the real in vivo situation. A set of test compounds with irritancy potencies especially in the mild and moderate range was tested. The improved LLBO showed some promising features which potentially could improve the usefulness of the BCOP test. Adaptation of cornea holders showed to be of limited value and only restricted to concentrations up to 15% which mimics more test conditions in industry.This 3-year research project was sponsored by the Stavros Niarchos Foundation (Greece).     

Effect of Hydration on Opacity in the Bovine Corneal Opacity and Permeability (BCOP) Assay

Abstract

The bovine corneal opacity and permeability (BCOP) assay has been proposed an an in vitro method for predicting the irritancy or toxicity of chemical substances. A loss of corneal transparency, which reduces visual acuity, can be the result of an increase in corneal hydration or direct damage to corneal tissues. The BCOP assay as currently practiced does not differentiate between these mechanisms. The purpose of this study was to determine the effect of increased hydration on transparency of the bovine cornea and to measure the hydration levels of corneas treated with compounds and substances known, from previous BCOP studies, to cause corneal opacity. Corneas were treated according to the BCOP protocol and corneal opacity was determined by measurement of light absorbance at 570 nm (A₅₇₀). Corneal hydration was determined by measurement of wet and dry weights. Hydration of the intact cornea was 3.86 mg H₂O/mg dry weight and A₅₇₀ = 0.048. After removal of the epithelium and incubation in MEM, corneal hydration increases to 6.27 mg H₂O/mg and A₅₇₀ = 0.11. Hydration of the maximally hydrated cornea, following removal of the epithelium and endothelium and incubation in deionized water, is 16.33 mg H₂O/mg and A₅₇₀ = 0.67. The hydration of corneas treated with compounds that are ranked as moderate to severe irritants based on the BCOP assay is not correlated with hydration levels of these corneas, since the A₅₇₀ of these corneas was usually higher than predicted from hydration levels. For example, after exposure to isopropanol hydration = 4.7, A₅₇₀ = 0.59; acetone: hydration = 5.69, A₅₇₀ = 1.38; 30% trichloroacetic acid: hydration = 4.28, A₅₇₀ = 1.43; 1% NaOH: hydration = 8.22, A₅₇₀ = 1.7. In contrast, after exposure to 30% sodium lauryl sulfate (SLS), which only removes epithelium, hydration = 5.38, A₅₇₀ = 0.095. This suggests that opacity in excess of that caused by increased hydration is caused by damage to corneal tissue, including epithelium and stroma. This was confirmed by light and transmission electron microscopy. Determining the cause of an increase in corneal opacity in the BCOP assay is important since tissue damage may be irreversible, while the cornea can recover from moderate increases in hydration. On the other hand, since increases in hydration alone cause relatively low A₅₇₀ readings, a potential irritant that causes a visually significant increase in cornea hydration might be ranked as a mild irritant. It is therefore proposed that measurement of corneal hydration be added to the BCOP protocol.     

Survey of ocular irritation predictive capacity using Chorioallantoic Membrane Vascular Assay (CAMVA) and Bovine Corneal Opacity and Permeability (BCOP) test historical data for 319 personal care products over fourteen years

The Chorioallantoic Membrane Vascular Assay (CAMVA) and Bovine Corneal Opacity and Permeability (BCOP) test are widely used to predict ocular irritation potential for consumer-use products. These in vitro assays do not require live animals, produce reliable predictive data for defined applicability domains compared to the Draize rabbit eye test, and are rapid and inexpensive. Data from 304 CAMVA and/or BCOP studies (319 formulations) were surveyed to determine the feasibility of predicting ocular irritation potential for various formulations. Hair shampoos, skin cleansers, and ethanol-based hair styling sprays were repeatedly predicted to be ocular irritants (accuracy rate = 0.90-1.00), with skin cleanser and hair shampoo irritation largely dependent on surfactant species and concentration. Conversely, skin lotions/moisturizers and hair styling gels/lotions were repeatedly predicted to be non-irritants (accuracy rate = 0.92 and 0.82, respectively). For hair shampoos, ethanol-based hair stylers, skin cleansers, and skin lotions/moisturizers, future ocular irritation testing (i.e., CAMVA/BCOP) can be nearly eliminated if new formulations are systematically compared to those previously tested using a defined decision tree. For other tested product categories, new formulations should continue to be evaluated in CAMVA/BCOP for ocular irritation potential because either the historical data exhibit significant variability (hair conditioners and mousses) or the historical sample size is too small to permit definitive conclusions (deodorants, make-up removers, massage oils, facial masks, body sprays, and other hair styling products). All decision tree conclusions should be made within a conservative weight-of-evidence context, considering the reported limitations of the BCOP test for alcohols, ketones, and solids.     

BioOcularTM角膜上皮模型ET50法在化妆品体外眼刺激评价中的应用

目的 采用国内生产的BioOcular?三维人源重组角膜上皮模型,对市面常见的几种洗发水和面霜的眼刺激性进行比较,通过该方法评价不同配方的化妆品在眼刺激性上的强弱。方法 采用三维人源重组角膜上皮模型BioOcular?模型 ET50法对不同品牌洗发水和面霜的眼刺激性进行比较研究,区分不同配方产品的眼刺激性强弱等级。结果 在同样实验条件下,洗发水和面霜表现出对三维人源重组角膜不同的刺激程度,不同配方品牌的洗发水的刺激程度也有很大差别。该方法可对化妆品的眼刺激性强弱程度进行很好的区分。结论 采用三维人源重组角膜上皮模型的ET50法是一种评价不同配方化妆品以及原料的眼刺激性的差异的快速替代检测方案。     

红霉素眼膏微生物限度检查方法的研究

建立红霉素眼膏微生物限度检查方法.利用红霉素在碱性条件下易分解失效的特点,采取在一定温度下调节pH值,对培养基进行稀释的方法建立红霉素眼膏微生物限度检查方法.红霉素眼膏的抗菌活性灭活后可有效检出污染菌.该方法可行性强,能有效地对红霉素眼膏进行微生物限度的测定.