N,N-diethylphenylacetamide, an insect repellent: absence of mutagenic response in the in vitro Ames test and in vivo mouse micronucleus test

N,N-diethylphenylacetamide (DEPA), a promising new insect repellent, was tested for mutagenicity in the in vitro Ames Salmonella/microsome mutagenicity test and the in vivo mouse micronucleus test. For the Ames test, DEPA was assayed both in the presence and absence of Aroclor 1254-induced rat-liver S-9 mix (5 and 20% S-9 fraction), using five tester strains of Salmonella typhimurium--TA97a, TA98, TA100, TA102 and TA104. For the micronucleus test, mice were exposed to DEPA through ip injection for 2 and 5 days in separate experiments, and bone marrow and peripheral blood were sampled 6 and 48 hr after the final injection, respectively. DEPA did not induce a mutagenic response in the Ames test, and mouse bone marrow and peripheral blood micronucleus tests. DEPA was not considered cytotoxic, as a depression of the percentage PCE was not observed at any dose in the range of 1 to 100 mg/kg body weight with either treatment protocol of the micronucleus test.     

An optimized method for the simultaneous determination of vitamins B1, B6, B12 in multivitamin tablets by high performance liquid chromatography

A simple, precise, rapid and selective HPLC-RP method has been developed for the simultaneous determination of thiamine hydrochloride (B(1)) 150 mg, pyridoxine hydrochloride (B(6)) 150 mg, and hydroxocobalamine chloride (B(12)) 0.150 mg in multivitamin tablets. The method uses a Hypersil-BDS C(18) reversed phase column and gradient elution. The aqueous mobile phase contained 0.015% triethylamine adjusted to pH 2.7 with 1 N sulfuric acid and acetonitrile. Separation and quantitation was achieved by changing the proportion of the system linearly with a time-schedule programme. Detection was carried out using a dual-beam UV detector set at 280, 350 nm. Good linearity was observed between the concentration of the analytes and peak area (r=0.9999, 0.9998). Sample preparation was relatively simple whereas excipients present in the dosage forms did not interfere with the peaks of interest. Recovery of the compound from the B(1), B(6), B(12) was quantitative.     

维生素B1、维生素B12肌注致过敏性休克

患者女,68岁,因右下肢酸痛半月余,于1999年9月23日来诊。经上级医院会诊后诊断为坐骨神经痛。于当日上午10时,给予维生素B_1 100mg、维生素B_(12)100μg,肌注。注射后约2min,患者突然出现恶心、胸闷、憋气、头晕、出冷汗、呼吸急促等症状。急扶患者平卧,查BP 30/0mmHg,患者面     

遗传毒性评价Ames试验方法的比较分析

Ames试验是一项体外致突变性试验,被广泛用于药物、食品、化学品和农药的遗传毒性检测,以评价受试物致突变的可能性。药物、食品、化学品和农药的指导原则和国家标准中Ames试验方法不尽相同,就这几者Ames试验方法的主要异同点进行比较分析,以期能熟练掌握Ames试验实施要点,更加规范实施检测工作,不断提高试验质量。     

Rescuing hepatocytes from iron-catalyzed oxidative stress using vitamins B1 and B6

In the following rescue experiments, iron-mediated hepatocyte oxidative stress cytotoxicity was found to be prevented if vitamin B1 or B6 was added 1 h after treatment with iron. The role of iron in catalyzing Fenton-mediated oxidative damage has been implicated in iron overload genetic diseases, carcinogenesis (colon cancer), Alzheimer’s disease and complications associated with the metabolic syndrome through the generation of reactive oxygen species (ROS). The objectives of this study were to interpret the cytotoxic mechanisms and intracellular targets of oxidative stress using “accelerated cytotoxicity mechanism screening” techniques (ACMS) and to evaluate the rescue strategies of vitamins B1 and B6. Significant cytoprotection by antioxidants or ROS scavengers indicated that iron-mediated cytotoxicity could be attributed to reactive oxygen species. Of the B6 vitamers, pyridoxal was best at rescuing hepatocytes from iron-catalyzed lipid peroxidation (LPO), protein oxidation, and DNA damage, while pyridoxamine manifested greatest protection against ROS-mediated damage. Thiamin (B1) decreased LPO, mitochondrial and protein damage and DNA oxidation. Together, these results indicate that added B1 and B6 vitamins protect against the multiple targets of iron-catalyzed oxidative damage in hepatocytes. This study provides insight into the search for multi-targeted natural therapies to slow or retard the progression of diseases associated with Fenton-mediated oxidative damage.     

Evaluation of genotoxicity of Antrodia cinnamomea in the Ames test and the in vitro chromosomal aberration test

Antrodia cinnamomea is an expensive and highly valued folk medicinal fungus that grows only inside the rotten trunk of Cinnamomum kanehirae, an evergreen broad-leaved tree. This fungus has recently been used commercially in the formulation of nutraceuticals and functional foods in Taiwan. It has been used for centuries as a detoxificant in cases of food poisoning, diarrhea, vomiting, hepatic disease and various kinds of cancers. The present study investigated the effects of Antrodia cinnamomea on mutagenicity using a bacterial reverse mutation assay employing the Salmonella typhimurium strains TA97, TA98, TA100, TA102, and TA1535. The effects of Antrodia cinnamomea on chromosome structure were tested in Chinese hamster ovary (CHO) cells. Antrodia cinnamomea was not mutagenic in all bacterial strains and it was not genotoxic in CHO cells.     

Rapid resolution liquid chromatography method development and validation for simultaneous determination of homocysteine,vitamins B6, B9, and B12 in human serum

Aim:

Hyperhomocysteinemia and vitamins B₆, B₉, and B₁₂ deficiencies usually result in various neurological, vascular, ocular, renal, and pulmonary abnormalities. However, to date, there are no simultaneous detection methods available for determining homocysteine, vitamins B₆, B₉, and B₁₂ levels in various biological fluids. In this study, we aim to develop a new validated simultaneous detection method for all four compounds to save both cost and time of analysis.

Materials and Methods:

The mobile phase consisted of a mixture of methanol and 1-heptanesulfonic acid sodium salt (33:67) with 0.05% triethylamine. The pH of the entire mixture was adjusted to 2.3 and the flow rate was 0.5 mL/min. Separation was achieved using a C-18 column (5 μm; 150 mm × 4.6 mm) maintained at 28°C in a column oven and the detection was conducted at 210 nm.

Results:

The method was linear between 50 and 1600 ng/mL for all of the drugs. The limits of detection for homocysteine, vitamins B₆, B₉, and B₁₂ were 5, 5, 10, and 10 ng/mL, respectively, while the limits of quantification were 10, 10, 25, and 25 ng/L, respectively. The developed method achieved good precision and accuracy and complies with the Food and Drug Administration (FDA) requirements.

Conclusion:

The developed and validated method is suitable to be used for the routine analysis of homocysteine, vitamins B₆, B₉, and B₁₂ simultaneously in human serum.KEY WORDS: Homocysteine, high performance liquid chromatography, simultaneous detection, vitamin B₆, vitamin B₉, vitamin B₁₂     

庆大维B胶囊中维生素B1、B2、B6和B12的HPLC法测定

建立HPLC法测定庆大维B胶囊中维生素B1、B2、B6和B12的含量。采用C18柱，乙腈-10mmol／L磷酸二氢钾（pH3．2）为流动相，梯度洗脱，检测波长280nm。维生素B1、B2、B6和B12分别在50～150、20-60、20～60和0.5～1．5μg/ml浓度范围内线性关系良好，平均回收率分别为101．9％、98．1％、99．6％和99．4％。     

Simultaneous determination of vitamins B1, B2, B6, and niacinamide in multivitamin pharmaceutical preparations by paired-ion reversed-phase high-pressure liquid chromatography

A high-pressure liquid chromatographic procedure for the simultaneous determination of vitamins B1, B2, B6, and niacinamide in multivitamin pharmaceutical preparations was developed and evaluated. The method uses paired-ion reversed-phase partition chromatography for baseline separation of the four water-soluble vitamins. This method was applied to the analysis of a multivitamin and multivitamin-multimineral tablets, and a technique was developed to reduce vitamin adsorption by the minerals. The results obtained by this method were compared with those obtained by the official methods. It was concluded that this method is fast, accurate, specific, and suitable for routine quality control use.     

Macrocyclic musk compounds--an absence of genotoxicity in the Ames test and the in vivo Micronucleus assay

Three synthetic macrocyclic musks, ethylene dodecanedioate, ethylene brassylate, and cyclopentadecanolide, which are widely used as ingredients of perfume fragrances, were tested for genotoxicity. In this report we present results from two different studies, the flow-cytometer-based micronucleus assay in peripheral blood of mice and the Salmonella/microsome test with TA 97, TA 98, and TA 100. Female NMRI and male CD 1 mice were intraperitoneally injected with one of the three macrocyclic musk compounds. Three different doses (0.1-1.6 g/kg bw) of each of the compounds were tested. Blood samples were collected on two occasions from each mouse and the frequency of micronucleated polychromatic erythrocytes (fMPCE) was determined. Neither of the compounds caused a significant difference from the control fMPCE. No mutagenic effect with and without S9 mix in the tested Salmonella strains was observed. The presence of S9 mix reduced the killing effect of high doses.     

Genotoxicity evaluation of titanium dioxide nanoparticles using the Ames test and Comet assay

Titanium dioxide nanoparticles (TiO₂‐NPs) are being used increasingly for various industrial and consumer products, including cosmetics and sunscreens because of their photoactive properties. Therefore, the toxicity of TiO₂‐NPs needs to be thoroughly understood. In the present study, the genotoxicity of 10nm uncoated sphere TiO₂‐NPs with an anatase crystalline structure, which has been well characterized in a previous study, was assessed using the Salmonella reverse mutation assay (Ames test) and the single‐cell gel electrophoresis (Comet) assay. For the Ames test, Salmonella strains TA102, TA100, TA1537, TA98 and TA1535 were preincubated with eight different concentrations of the TiO₂‐NPs for 4 h at 37 °C, ranging from 0 to 4915.2 µg per plate. No mutation induction was found. Analyses with transmission electron microscopy (TEM) and energy‐dispersive X‐ray spectroscopy (EDS) showed that the TiO₂‐NPs were not able to enter the bacterial cell. For the Comet assay, TK6 cells were treated with 0–200 µg ml^–1 TiO₂‐NPs for 24 h at 37 °C to detect DNA damage. Although the TK6 cells did take up TiO₂‐NPs, no significant induction of DNA breakage or oxidative DNA damage was observed in the treated cells using the standard alkaline Comet assay and the endonuclease III (EndoIII) and human 8‐hydroxyguanine DNA‐glycosylase (hOGG1)‐modified Comet assay, respectively. These results suggest that TiO₂‐NPs are not genotoxic under the conditions of the Ames test and Comet assay. Published 2012. This article is a US Government work and is in the public domain in the USA.     

Neratinib reverses ATP-binding cassette B1-mediated chemotherapeutic drug resistance in vitro, in vivo, and ex vivo

Neratinib, an irreversible inhibitor of epidermal growth factor receptor and human epidermal receptor 2, is in phase III clinical trials for patients with human epidermal receptor 2-positive, locally advanced or metastatic breast cancer. The objective of this study was to explore the ability of neratinib to reverse tumor multidrug resistance attributable to overexpression of ATP-binding cassette (ABC) transporters. Our results showed that neratinib remarkably enhanced the sensitivity of ABCB1-overexpressing cells to ABCB1 substrates. It is noteworthy that neratinib augmented the effect of chemotherapeutic agents in inhibiting the growth of ABCB1-overexpressing primary leukemia blasts and KBv200 cell xenografts in nude mice. Furthermore, neratinib increased doxorubicin accumulation in ABCB1-overexpressing cell lines and Rhodamine 123 accumulation in ABCB1-overexpressing cell lines and primary leukemia blasts. Neratinib stimulated the ATPase activity of ABCB1 at low concentrations but inhibited it at high concentrations. Likewise, neratinib inhibited the photolabeling of ABCB1 with [(125)I]iodoarylazidoprazosin in a concentration-dependent manner (IC(50) = 0.24 μM). Neither the expression of ABCB1 at the mRNA and protein levels nor the phosphorylation of Akt was affected by neratinib at reversal concentrations. Docking simulation results were consistent with the binding conformation of neratinib within the large cavity of the transmembrane region of ABCB1, which provides computational support for the cross-reactivity of tyrosine kinase inhibitors with human ABCB1. In conclusion, neratinib can reverse ABCB1-mediated multidrug resistance in vitro, ex vivo, and in vivo by inhibiting its transport function.     

Some interactions of light, riboflavin, and aflatoxin B1 in vivo and in vitro.

In previous studies, artificial sunlight and riboflavin synergistically increased acute aflatoxin toxicity in rats. Three new experiments were designed to provide information on the interaction of riboflavin, aflatoxin, and light. In a study of carcinogenesis, rats received low levels of aflatoxin 5 days/wk for 3 wk; 30 min after each dosing, half of them were irradiated for 2 hr. In some, levels of glucose-6-phosphatase and acid phosphatase were determined 5 days after completion of treatment. Remaining rats were killed at 30 or 53 wk. All underwent complete necropsies and histopathologic examination. In the second experiment, rats were dosed with riboflavin and divided into four groups: no further treatment; aflatoxin (LD50); irradiation (1-2 hr); or aflatoxin plus irradiation. Blood riboflavin levels were determined at intervals following these treatments. In the third experiment, the chemical reactions of irradated aflatoxin and/or riboflavin were studied by uv spectroscopy and TLC. The 53-wk study showed clearly that light decreased the incidence of aflatoxin-induced cancer. The other results may provide an explanation. Aflatoxin caused blood riboflavin levels to decrease-an effect enhanced by irradiation, suggesting that photosensitized riboflavin and aflatoxin form a complex. This interpretation gains support from studies in vitro that showed that riboflavin quenched aflatoxin photodegradation, perhaps by complexing with aflatoxin. Thus, low, carcinogenic doses of aflatoxin may complex with endogenous, photosensitized riboflavin, inhibiting its degradation into carcinogenic metabolites.     

Use of the Ames test in toxicology

The Salmonella/microsome assay (Ames test) is one of the most widely used short-term tests. Despite the ubiquitous presence of this assay and the large number of chemicals tested, there is still controversy over the value of Salmonella/microsome assay results in risk assessment. Understanding of the properties and performance of the test system is necessary to provide an appropriate answer to this question. Based on both theoretical and empirical considerations, the results of the assay can provide useful information to assist in the development of further studies and as part of the data which can be used in evaluating potential biological effects or projected lack of adverse effects. The value of the Salmonella/microsome assay in performing these functions is considerably enhanced by consideration of all the information which the assay data provide.     

Genotoxic evaluation of Halfenprox using the human peripheral lymphocyte micronucleus assay and the Ames test

The genotoxicity and mutagenicity of Halfenprox, a synthetic pyrethroid insecticide and acaricide, was assessed using two standard genotoxicity assays of the Salmonella typhimurium mutagenicity assay (Ames test) and in vitro micronucleus (MN) assay in human peripheral lymphocytes. In the Ames test, Salmonella strains TA98 and TA100 were treated with or without S9 fraction. The doses of Halfenprox were 6.25, 12.5, 25, 50, and 100?μg/plate and test materials were dissolved in DMSO. The concentrations of Halfenprox did not show mutagenic activity on both strains with and without S9 fraction. The MN assay was used to investigate the genotoxic effects of Halfenprox in human peripheral lymphocytes treated with 250, 500, 750, and 1000?μg/ml concentrations of Halfenprox for 24 and 48?h, and at 1000?μg/ml the concentration was significantly increased and the MN formation was compared with the negative control for both treatment periods. In addition, a significant decrease of the nuclear devision index (NDI) values at the higher concentrations of Halfenprox and at both treatment periods was observed.