Hemolysis of human erythrocytes induced by melamine-cyanurate complex

Melamine is a widely-used chemical in industries. In recent years, melamine has been found to be involved in outbreaks of renal injury in infants and animals. Pathological studies indicated that the melamine-induced acute renal failure was related to the concurrence of melamine and other triazine analogs such as cyanuric acid. In the present study, human erythrocytes were used as an in vitro model to explore the cytotoxicity of melamine and its complex with cyanuric acid. The results demonstrated that mixing melamine and cyanuric acid resulted in the formation of insoluble particles and that the insoluble melamine-cyanurate complex induced membrane damages of human erythrocytes. The membrane damages included hemolysis, K⁺ leakage, alterations in cell shape and membrane fragility, and inhibition of enzymatic activity. By contrast, either melamine or cyanuric acid alone had no effect on erythrocyte membranes. The results of this study may provide a fresh insight into the melamine toxicology.     

三聚氰胺对花鲈的急性毒性实验研究

本文以花鲈(Lateolabrax japonicus)为实验对象,采用接触、腹腔注射及口服三种致毒方式,进行了非蛋白氮物质-三聚氰胺对花鲈的急性毒性实验研究。口服急性毒性实验中,通过在饲料中添加不同浓度的三聚氰胺（0、500、2000、5000及10000 mg/kg）,进行了21d的花鲈养殖实验。实验结果表明：三聚氰胺溶解度较低,其水溶液没有表现出急性毒性,LC50 > 3500 mg/L;而在腹腔注射致毒方式下,三聚氰胺对花鲈的半致死剂量LD50 = 890.07 mg/kg•w;LD5095%可信限为：778.63－1017.45 mg/kg•w;在口服急性毒性实验中,10000 mg/kg组三聚氰胺降低了花鲈的摄食与生长(p<0.05),饲料系数显著升高 (p<0.05);三聚氰胺对花鲈的存活率、肥满度、肝体比及脏体比均无显著影响 (p>0.05)。饲料中添加三聚氰胺没有显著影响花鲈血清的丙氨酸氨基转移酶、天冬氨酸氨基转移酶、总蛋白、葡萄糖、胆固醇、甘油三酯及尿素氮 (p>0.05),但显著影响了花鲈血清的碱性磷酸酶活性,10000 mg/kg组的碱性磷酸酶活性显著高于其他各处理组 (p<0.05),其他各组间无显著差异(p>0.05)。三聚氰胺对花鲈21d的最大未观察到有害作用剂量（NOAEL, no-observed-adverse-effect-level）为131.99 mg/kg•w•d。     

Spectroscopic investigation on the toxic interaction of melamine with herring sperm DNA

The toxic interaction of melamine with herring sperm DNA (hs‐DNA) was investigated by using fluorescence and UV–vis absorption spectra techniques. The experimental results showed that the toxic interaction between melamine and hs‐DNA occurred. Fluorescence quenching experiments indicated the existence of electrostatic binding between melamine and hs‐DNA. The binding constants K_A and the binding site numbers were calculated by means of the Stern–Volmer equation and were 9.8 × 10⁴ L mol^?1 and 1.3, respectively. Both the results of fluorescence spectra and UV–vis absorption spectra verified that there are electrostatic binding between melamine and hs‐DNA. The possibility in the presence of a classical intercalation binding mode could be ruled out by using DNA unwinding experiments. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 24:323–329, 2010; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.20341     

高浓度三聚氰胺对萼花臂尾轮虫 生殖策略的影响

以萼花臂尾轮虫(Brachionus calyciflorus)为受试动物,研究了不同浓度(400、800和1 200 mg/L)的三聚氰胺对萼花臂尾轮虫种群增长、有性生殖、个体大小和卵大小的影响.结果显示,三聚氰胺对萼花臂尾轮虫的24 h LC50值为2 627.00 mg/L.与空白对照组相比,800和1 200 ...     

Molecularly-imprinted microspheres for selective extraction and determination of melamine in milk and feed using gas chromatography–mass spectrometry

Molecularly-imprinted polymers in the form of microspheres were synthesized using the dispersion polymerization protocol; cyromazine was used as dummy template, while methacrylic acid, ethylene glycol dimethacrylate and acetonitrile (MeCN) were used as functional monomer, cross-linker, and porogen, respectively. When compared with the non-imprinted polymer, the molecularly-imprinted polymers (MIPs) showed outstanding affinity toward melamine in MeCN with a maximum binding concentration (B_max) of 53.20 nmol mg⁻¹ MIPs, imprinting effect of 4.6, and a dissociation constant (K_d) of 90.45 μM. After optimization of the molecularly-imprinted solid-phase extraction conditions, a new method was developed to determine the melamine in milk and feed with gas chromatography–mass spectrometry. The performance of this method has been evaluated in the tainted milk and feed in terms of recovery, precision, linearity, the limit of detection (LOD) and limit of quantitation (LOQ). Recovery ranged in samples from 93.1 to 101.3% with intra-day and inter-day relative standard deviation values below 5.34%. The LOD and LOQ of melamine in milk and feed were 0.01 μg mL⁻¹ (μg g⁻¹) and 0.05 μg mL⁻¹ (μg g⁻¹), respectively.     

Effects of Melamine and Cyanuric Acid on Embryo‐Fetal Development in Rats

After the outbreak of acute renal failure associated with melamine‐contaminated pet food, melamine and melamine‐related compounds have become of great interest from a toxicologic perspective. We investigated the potential effects of melamine in combination with cyanuric acid (M + CA, 1:1) on pregnant dams and embryo‐fetal development in rats. M + CA was orally administered to pregnant rats from gestational days 6 through 19 at doses of 0, 3, 10, and 30 mg/kg/day of both melamine and cyanuric acid. Maternal toxicity of rats administered 30 mg/kg/day M + CA was manifested as increased incidences of clinical signs and death; gross pathologic findings; higher blood urea nitrogen and creatinine levels; lower body weight gain and food intake; decreased thymus weight; and increased heart, lung, and kidney weights. Histopathological examinations revealed an increase in the incidence of congestion, tubular necrosis/degeneration, crystals, casts, mineralization, inflammatory cells in tubules, tubular dilation, and atrophy of glomeruli in maternal kidneys, whereas fetal kidneys did not show any histopathological changes. Developmental toxicity included a decrease in fetal (28%) and placental weights and a delay in fetal ossification (n = 7). Increased incidence of gross and histopathological changes in the maternal kidney was also found in the middle dose group (n = 12). No treatment‐related maternal or developmental effects were observed in the low dose group (n = 12). Under these experimental conditions, M + CA is embryotoxic at an overt maternotoxic dose in rats and the no‐observed‐adverse‐effect level of M + CA is considered to be 3 mg/kg/day for pregnant dams and 10 mg/kg/day for embryo‐fetal development.     

Colorimetric detection of melamine in milk by citrate-stabilized gold nanoparticles

Here, we report a simple and sensitive colorimetric method for detection of melamine in milk using gold nanoparticles (AuNPs). AuNPs of 21-nm size were synthesized by the citrate reduction method. The method is based on the principle that the melamine causes the aggregation of AuNPs and, hence, the wine red color of AuNPs changes to blue or purple. This change in color can be visualized with the naked eye or an ultraviolet–visible (UV–Vis) spectrometer. Under optimized conditions, AuNPs are highly specific for melamine and can detect melamine down to a concentration of 0.05 mg L⁻¹.     

A novel mixed-mode solid phase extraction for simultaneous determination of melamine and cyanuric acid in food by hydrophilic interaction chromatography coupled to tandem mass chromatography

Utilizing a solid phase extraction column (MCT) containing mixed hydrophilic functional gel and cation exchange sorbent, a sensitive and rapid HPLC–MS/MS method for simultaneously determining the residues of melamine (MEL) and cyanuric acid (CYA) in human foodstuffs was developed. MEL and CYA in egg, pork, liver, kidney and pork, shrimp, sausage casing, honey, soybean milk, soybean powder and dairy product were extracted using acetonitrile/water, defatted with hexane and isolated using MCT solid phase extraction column. The residues were separated upon a hydrophilic interaction liquid chromatography (HILIC) column and analyzed by electrospray ionization under negative–positive switched mode on a triplequadrupole mass spectrometry. The selected reaction monitoring was performed on [M+H]⁺ of m/z 127.9 to provide the transition of 127 > 85 and 127 > 68 (MEL) while the [M−H]⁻ of m/z 127.1 was selected as the precursor ion for CYA resulting in product ions m/z 85 and 42. Isotope labeled internal standard (¹⁵N₃-MEL and ¹³C₃-CYA) and matrix-matched calibration were both used to observe the recovery to be 70.0–129.6% and 70.0–128.9% with RSD of 1.4–23.3% and 1.5–21.7% for MEL and CYA, respectively (n = 6). All the LODs and LOQs of MEL and CYA were less than 39.4 and 99.1 μg kg⁻¹, respectively, in 18 matrices, which were sensitive enough for quantitative analysis. This method has been proven effective in simultaneous determination of melamine and cyanuric acid when inspecting unknown and positive samples.     

Smartphone‐based portable wireless optical system for the detection of target analytes

Rapid and accurate on‐site wireless measurement of hazardous molecules or biomarkers is one of the biggest challenges in nanobiotechnology. A novel smartphone‐based Portable and Wireless Optical System (PAWS) for rapid, quantitative, and on‐site analysis of target analytes is described. As a proof‐of‐concept, we employed gold nanoparticles (GNP) and an enzyme, horse radish peroxidase (HRP), to generate colorimetric signals in response to two model target molecules, melamine and hydrogen peroxide, respectively. The colorimetric signal produced by the presence of the target molecules is converted to an electrical signal by the inbuilt electronic circuit of the device. The converted electrical signal is then measured wirelessly via multimeter in the smartphone which processes the data and displays the results, including the concentration of analytes and its significance. This handheld device has great potential as a programmable and miniaturized platform to achieve rapid and on‐site detection of various analytes in a point‐of‐care testing (POCT) manner.     

三聚氰胺等物质给予幼年恒河猴的毒性试验

目的观察三聚氰胺及其同系物给予动物后,导致肾脏损伤、形成结石的过程,及肾脏损伤的恢复过程。方法选择6月龄恒河猴6只,雌雄各半,随机分为3组,2只/组,分别为T1（140 mg/kg三聚氰胺和14 mg/kg三聚氰酸）,T2（140 mg/kg三聚氰胺和50mg/kg腺嘌呤）和T3组（140mg/kg三聚氰胺）。连续70 d口服受试物,观察动物的一般状态,分别于不同时间点进行血液学、血生化、尿常规和肾脏B超的检测;并连续观察18个月,观察动物的生长发育情况。结果 1.各剂量组动物在给予受试物8周后所有动物肾脏中均出现多个结石;停止给予受试物后,所有动物结石数量和大小均呈下降趋势,至停止给予受试物16个月后,全部动物肾脏均未检测出结石。2.各剂量组,血清中β2微球蛋白（β2-MG）和尿酸（UA）等指标随着给药时间的延长而有所增高,但在正常范围内。在动物的恢复期,此两项指标的数值明显下降。结论 1.单纯三聚氰胺（相当于人体使用量48 mg/kg.w.d）不能造成恒河猴肾脏功能的明显损害,可形成肾结石。2.在三聚氰酸或腺嘌呤存在的情况下,三聚氰胺可以造成恒河猴肾功能指标β2-MG和UA在正常范围内升高,并可在肾脏形成结石;。3.经过16个月的恢复所有动物结石无法检出,其中三聚氰胺组动物恢复最快。