植物微核技术在监测水质污染中的应用

应用韭菜微核技术,检测哈尔滨市量具刃具厂含铬废水的遗传毒性,试验表明：含铬废水可诱导韭菜和蚕豆根尖细胞产生微核,进行回归分析,其蚕豆对Ｃｒ＾＋６诱变剂回归分析为ｙ＝２４．９４＋１２７．７７ｘ（ｒ＝０．９９６,Ｐ〈０．０１）,韭菜对Ｃｒ＾＋６诱变剂回归广泛为：ｙ＝２１．７４＋１００．２３ｘ（ｒ＝０．９００,Ｐ〈０．０１）,ｒ＝１１．１７＋３．３８ｘ（ｒ＝０．７８１,Ｐ〈０．０５）,韭菜根尖微核技术具     

Evaluation of the Allium anaphase-telophase test in relation to genotoxicity screening of industrial wastewater

The Allium anaphase-telophase test was evaluated to find out if it could be recommended in the screening of wastewater for genotoxicity. Five mutagenic or carcinogenic chemicals usually found in wastewater were tested in the Allium anaphase-telophase test. Sodium dichromate (25 μM), benzene (100 μM), dichloromethane (175 μM) and 1,1,1-trichloromethane (175 μM) increased the frequency of chromosome aberrations in the root cells, whereas formaldehyde (1 mM) was found to be non-mutagenic in this test system. Other studies where chemicals were tested in the Allium test were reviewed. For 15 chemicals the results were compared with results from the Ames test, the Microscreen assay, and carcinogenicity tests in rodents. The sensitivity of the Allium test was calculated to be 82%. In conclusion the Allium test is recommended for the screening of wastewater because it has a high sensitivity, is cheap, rapid, easy to handle, and because it can be used on wastewater without pretreatment of the sample.     

Biomonitoring of low levels of mercurial derivatives in water and soil by Allium micronucleus assay

The Allium micronucleus (MNC) assay was developed to monitor low levels of mercury in aquatic and terrestrial environments. Four mercurial derivatives namely mercuric chloride (MC), methyl mercuric chloride (MMC), phenyl mercuric acetate (PMA) and a methoxy ethyl mercuric chloride based fungicide, Emisan-6, were tested to assess the sensitivity and versatility of the Allium MNC assay. Allium bulbs were set directly on water and soil contaminated with known levels of mercurial derivatives (0.0001-10.00 ppm). On the 5th day the endpoints measured were root length, mitoses with spindle abnormality and cells with MNC in root meristems. The effective concentrations of the test chemicals that cause 50% of root length as compared to control (EC50) were determined from dose-response curves so obtained. The lowest effective concentration tested (LECT) and highest ineffective concentration tested (HICT) for each of the mercurial derivatives for the induction of spindle malfunction and MNC were determined. It was found that EC50, LECT and HICT values for mercurial derivatives in soil were higher than those in water. The frequencies of cells with MNC and mitoses with spindle abnormality were highly correlated indicating that MNC is a good parameter of spindle malfunction. The present approach increased the sensitivity of the Allium assay by 10-fold, the detection limit being 0.001-0.1 ppm and 0.1-1.0 ppm in aquatic and terrestrial environments respectively, depending on the species of mercury.     

Genotoxicity of industrial wastewaters obtained from two different pollution sources in northern India: a comparison of three bioassays

The genotoxicity of industrial wastewater samples from Aligarh and Ghaziabad cities was compared using the Ames plate incorporation test, the Ames fluctuation test and the Allium cepa test. While TA102 and TA104 strains exhibited the highest sensitivity against the Aligarh sample (AWW) in terms of the slope (m) of the dose-response curve in the plate incorporation assay, TA98 and TA97a were the most sensitive strains based on the induction factor, Mi(p). TA98 once again, was the most sensitive strain against the test sample from Ghaziabad (GWW) in terms of 'Mi(p)' while TA102 was the most sensitive strain on the basis of the slope (m). TA100 displayed the highest susceptibility towards the samples from Aligarh in the fluctuation test. However, TA102 and TA98 responded maximally to GWW in this bioassay. The mutagenicity of the test samples seemed to be partly mediated by reactive oxygen species (ROS) as evidenced by the use of free radical scavengers. Mannitol brought about the maximum decline in the number of revertants of TA102 by the Aligarh sample, whereas such a reduction in case of Ghaziabad sample was exhibited with superoxide dismutase. Both the test water samples induced various anaphase aberrations in the root cells of Allium cepa. Fragmentation of the chromosome was the predominant effect of the Aligarh water sample while the Ghaziabad sample induced chromosome stickiness. The crucial roles of heavy metals and pesticides in the genotoxicity of AWW and GWW, respectively, have been suggested. In view of the problem associated with the interpretations of data, we recommend that all the test bioassays should be carried out in the presence of ROS scavengers for the fool proof evaluation of the genotoxicity of water samples.     

The improved Allium/Vicia root tip micronucleus assay for clastogenicity of environmental pollutants

The meristematic mitotic cells of plant roots are appropriate and efficient cytogenetic materials for the detection of clastogenicity of environmental pollutants, especially for in situ monitoring of water contaminants. Among several cytological endpoints in these fast dividing cells, such as chromosome/chromatid aberrations, sister-chromatid exchanges and micronuclei, the most effective and simplest indicator of cytological damage is micronucleus formation. Although the Allium cepa and Vicia faba root meristem micronucleus assays (Allium/Vicia root MCN) have been used in clastogenicity studies about 12 times by various authors in the last 25 years, there is no report on the comparison of the efficiency of these two plant systems and in different cell populations (meristem and F₁) of the root tip as well as under adequate recovery duration. In order to maximize the efficiency of these bioassays, the current study was designed to compare the Allium and the Vicia root MCN assays on the basis if chromosome length, peak sensitivity of the mitotic cells, and the regions of the root tip where the MCN are formed. The total length of the 2n complement of Allium chromosomes is 14.4 μm and the total length of the 2n complement of Vicia is 9.32 μm. The peak sensitivity determined by serial fixation at 12-h intervals after 100 R of X-irradiation is 44 h. The slope of the X-ray dose-response curve of Allium roots derived from the meristematic regions was lower than that derived from cells in the F₁ region. Higher efficiency was also demonstrated when the MCN frequencies were scored from the F₁ cells in both Allium and Vicia treated with formaldehyde (FA), mitonycin C (MMC), and maleic hydrazide (MH). The results indicated that scoring of MCN frequencies from the F₁ cell region of the root tip was more efficient than scoring from the meristematic region. The X-ray linear regression dose-response curves were established in both Allium and Vicia cell systems and the coefficients of correlations, slope values were used to verify the reliability and efficiency of these two plant cell systems. Based on the dose-response slope value of 0.894 for Allium and 0.643 for Vicia, the Allium root MCN was a more efficient test system. The greater sensitivity of the Allium roots is probably due to the greater total length of the diploid complement and the higher number of metacentric chromosomes. The Allium/Vicia root MCN test system was applied to determine the clastogenicity of saccharin (SC) and wastewater from Rio Queretaro and the Arena canal in the city of Queretaro, Mexico. The minimum effective dose (MED) is 10 R for X-rays, 50 mM for FA, 2.2 mM for MMC, 0.01 mM for MH, and 40 ppm for SC.     

Cytotoxic and genotoxic effects of dioxacarb by human peripheral blood lymphocytes CAs and Allium test

Dioxacarb (Elecron, Famid) is a phenyl methylcarbamate insecticide and in vitro cytotoxic and genotoxic effects of this pesticide on human peripheral blood lymphocytes and Allium root meristematic cells were investigated by chromosomal aberrations (CAs) and Allium test. Human lymphocytes were treated with 62.5, 125, 250 and 500 ppm doses of dioxacarb for CAs. CA/cell, abnormal cell % and mitotic index % (MI %) data were obtained from these concentrations in 24 and 48 h treatment periods. Dioxacarb did not increase the CA/cell frequency significantly, so this insecticide was not identified as genotoxic. But it was found cytotoxic especially at 250 and 500 ppm concentrations because of the reduced the MI % and increased the abnormal cell %. In Allium test, 25 ppm (EC50/2), 50 ppm (EC50) and 100 ppm (EC50 × 2) concentrations were used for root growth inhibition (EC50 determination) and Allium mitotic index (MI) determination tests. The used concentrations of dioxacarb induced dose-dependent inhibition of MI and root growth on root meristems. Mitotic inhibition of dioxacarb was found significantly higher than for the positive control. These Allium results indicated the high cytotoxicity of dioxacarb. The present study is the first research on cytotoxicity and genotoxicity of dioxacarb by human lymphocyte CAs and Allium test.     

沙葱种皮特性、种胚及种子浸提液与种子休眠的关系

本文以沙葱休眠种子为试材,检测其种皮透水性、透气性、超微结构、种胚休眠及种子浸提液。结果表明,酸蚀种子吸水12h达饱和,吸水率是完整种子的2倍;其呼吸强度最大值是完整种子的1.36倍。酸蚀处理使种子表面结构破损,角质层脱落,种孔露出。沙葱种子离体胚接种培养后2～3d,种胚开始萌发,萌发率达85％。沙葱完整种子与粉碎种子的浸提液对白菜种子发芽率、苗高、根长均有抑制作用,且随着浓度的提高而增强。     

Genotoxicity biomarkers in occupational exposure to formaldehyde--the case of histopathology laboratories

Oxidative stress-mediated genotoxicity of wastewaters taken from two different cities, Saharanpur (SWW) and Aligarh (AWW), were compared with a battery of short-term assays namely the Allium cepa genotoxicity test, the plasmid-nicking assay, and the Ames fluctuation test. Both test-water samples - when used undiluted - increased the frequency of chromosomal abnormalities and/or micronuclei and alterations in the mitotic index of root cells of Allium cepa. Bridges and fragmentation of the chromosome were the predominant effects of the Saharanpur water sample while the Aligarh sample induced mainly chromosome fragmentation. Single- and double-strand breaks were also observed in plasmid DNA treated with these test wastewaters. The plasmid-nicking assay performed on SWW resulted in linearization of plasmid DNA when 18μl was tested (in a total reaction volume of 20μl). However, with the same amount of AWW, all three forms of plasmid, viz. supercoiled, open circular and linear were observed. Supplementation with specific scavengers of reactive oxygen species (ROS) caused a significant decline in mutagenicity of test-water samples in all the tests, pointing at oxidative stress as the mediator of the observed genotoxicity. The role of heavy metals in the AWW-induced oxidative stress and that of phenolics in SWW cannot be ruled out.     

Antifungal effects of the volatile oils from Allium plants against Trichophyton species and synergism of the oils with ketoconazole

In an attempt to develop stable and safe antifungal agents from natural products (daily foodstuffs in particular), the activities of essential oils from Allium sativum for. pekinense, A. cepa, and A. fistulosum against three Trichophyton species responsible for severe mycoses in humans were investigated and compared with activity of allicin in this study. The fungistatic activities of Allium oils were evaluated by the broth dilution method and disk diffusion assay. The combined effects of Allium oils with ketoconazole were tested by the checkerboard titer test. Among the tested oils, A. sativum for. pekinense oil exhibited the strongest inhibition of growth of T. rubrum, T. erinacei, and T. soudanense with MICs (minimum inhibiting concentrations) of 64microg/ml, while the activities of A. cepa and A. fistulosum were relatively mild. The inhibiting activities of the oils on Sabouraud agar plates were dose dependent against Trichophyton species. Additionally, these oils showed significant synergistic antifungal activity when combined with ketoconazole in the checkerboard titer test and disk diffusion test.     

A study of the migration and degradation of oil in an upland bog peat soil in the Khanty-Mansi autonomous district during biotesting

Water migration and the degradation of Shaim oil in soils of upland bogs in the Khanty-Mansi Autonomous District were studied under laboratory conditions and the toxicity of consecutive samples of water extracts was investigated using Ceriodaphnia affinis, Paramecium caudatum, and Allium sera as test objects. It was found that oil (up to 2 g/kg) is retained by the humic matrix of peat and does not pass into water. The biotesting method permits one to fix the toxic effect of leachate from oil-contaminated peat, even in cases when petroleum products are not detected by means of IR spectrometry.     

Genotoxicity of hydrated sulfur dioxide on root tips of Allium sativum and Vicia faba

Genotoxicity of sulfur dioxide (SO(2)) and its hydrates (bisulfite and sulfite) in human lymphocytes and other mammalian cells have been found earlier in our laboratory. In the present studies, we used Allium stavium and Vicia faba cytogenetic tests, which are the highly sensitive and simple plant bioassays. A mixture of sodium bisulfite and sodium sulfite (1:3), at various concentrations from 1 x 10(-4) to 2 x 10(-3)M was used for the treatment. Genotoxicity was expressed in terms of anaphase aberration (AA) frequencies in the Vicia-AA test and in terms of micronuclei (MCN) frequencies in both Vicia-MCN test and Alllium-MCN test. On average, the results showed a 1.7-3.9-fold increase of AA frequencies and a 3.5-4.5-fold increase of MCN frequencies in Vicia root tips as compared with the negative control. Similarly, results of Allium-MCN test also showed a significant increase in MCN frequencies in the treated samples. In addition, pycnotic cells (PNC) appeared in Allium root tips of treated groups. The frequencies of MCN, AA and PNC increased dose-dependently and the cell cycle delayed at the same time in bisulfite treated samples. Results of the present study suggest that the Vicia and Allium cytogenetic bioassays are efficient, simple and reproducible in genotoxicity studies of bisulfite.     

Inhibitory effect of adrenaline and hydrocortisone on the growth of Allium cepa roots

The roots of Allium cepa were allowed to grow in distilled water containing 10(-4) M adrenaline hydrochloride or 2 X 10(-3) M hydrocortisone sodium succinate. Adrenaline inhibited the growth of roots; they decreased in length, number and total dry weight. The total amount of DNA in the roots was reduced much less than that of extracellular root components after adrenaline. Also hydrocortisone treatment resulted in a considerable decrease of the length and dry weight of Allium cepa roots. Both DNA and extracellular root components were influenced.     

Changes of abscisic acid and auxin as related to dormancy breaking ofAllium wakegi bulblets by vacuum infiltration and BA treatment

Dormant bulblets of the sterile green onion,Allium wakegi, broke dormancy and sprouted when infiltrated with water for 1 h at the reduced pressure of 25 kPa or when dipped in 20 ppm of benzylaminopurine (BA) for 1 h at normal (100 kPa) or reduced pressure. One day after the treatment, the abscisic acid (ABA) contents greatly decreased in bulblets treated with BA at both reduced and normal pressure as well as in bulblets treated with water at reduced pressure. The ABA content of bulblets dipped only in water at normal pressure remained relatively high throughout the study. Changes in 2-trans-abscisic acid (t-ABA) and IAA content of bulblets showed no correlation with breaking of dormancy. Dormancy breaking ofAllium wakegi bulblets appears to be related to a sharp decrease in ABA content in the bulblets as a result of water infiltration or BA treatment.     

Phytotoxicity of colloidal solutions of metal-containing nanoparticles

Phytotoxicity of colloidal solutions of metal-containing nanoparticles (Ag, Cu, Fe, Zn, Mn) was investigated using a standard Allium cepa (L.) test system. Toxicity of experimental solutions at the organism level was evaluated in terms of biomass growth of onion roots, and cytotoxicity was estimated by proliferative activity of root meristem cells. The colloidal solutions of metal nanoparticles inhibited the growth of Allium cepa (L.) roots due to their ability to penetrate into cells and interact with their components and, thus, to inhibit mitosis. According to our results, cytotoxicity of test solutions decreases in the following order: Cu ≥ Zn ≥ Ag ≥ Fe. Solution of Mn-containing nanoparticles contributed to root growth reaction.     

Analysis of the cell cycle in the root meristem of Allium cepa under the influence of ledakrin

The influence of a polish anticancer drug on the cell cycle using Allium test was studied. Methods of aceto-orcein squash slides, curve of labelled mitoses after 3H-thymidine incubation and cytophotometrics after Feulgen's reaction were employed. Ledakrin acts strongly antimitotically, but it does not block the cell cycle completely. The cytostatic activity of ledakrin results from its action on the interphase. The phases G1 and S are prolonged while M is unchanged after 6h incubation with ledakrin. During postincubation in water without ledakrin it was noted, at the beginning, that the mitotic activity decreases and it is brought about the lengthening of S and G2 phases. The duration of the cell cycle phases returns to the control level during further postincubation. The results of analysis of chromatin aberrations and the micronucleus test point to a mutagenic effect of ledakrin.     

Induction of polyploidy and C-tumours after treatingAllium cepa root tips with the herbicide “Treflan”

Treflan has the ability to induce polyploidy inAllium cepa root tips. The frequency of polyploidy was reduced after allowing the roots to recover, which indicates that the process of polyploidy is a reversible one, if we apply Treflan for a short time (4 h). As soon as the chemioal is removed from the cells, they go on with their normal way in division. It was found that the concentration 350 × 10^-5 ml Treflan per 100 ml water is an effective concentration in producing polyploidy in the roots ofAllium cepa. Treflan induced C-tumours in root tips ofAllium cepa. Two types of enlargement were noticed, complete swelling along the majority of root tips and complete swelling with a certain enlargement immediately after 2 mm terminal tips. The histological examination of these zones revealed that swellings were due to cell enlargement both in meristematic and in tumour zones. This conclusion was evidenced by the results obtained from the nucleoplasmic index.     

Genetic effects of root extracts of Glycyrrhiza glabra L. in different test systems

The antimutagenic and geroprotective activities of licorice (Glycyrrhiza glabra) root extracts were established in cells of plant test systems of Allium fistulosum L., Allium cepa L., and Vicia faba L. and of laboratory animals (Vistar rats). The prospects for the practical application of licorice Glycyrrhiza glabra root extracts as antimutagenic substances are discussed.     

Relationships between induction of anesthesia and mitotic spindle disturbances studied by means of principal component analysis

A dataset comprising the activity of 30 compounds in 4 biological tests--anesthesia of tadpoles, anesthesia of frog heart, abnormal growth and spindle disturbances in Allium root tips--was re-evaluated by means of principal component analysis. A two-component model is required to explain the variation in biological activity of the compounds. It is found that abnormal growth is different from the other biological responses. When this test is excluded, as much as 90% of the variation is explained by a one-component model, the determining factor most probably being the lipophilic character of the compounds. Mammalian mitotic cells respond in a similar way to mitotic cells of Allium root tips. It is suggested that possible regularities in the dose-response relationships for anesthesia, teratogenic effects and generation of abnormal chromosome numbers require further exploration.     

响应面分析法优化大蒜蒜氨酸提取工艺

以新鲜大蒜为原料,微波灭酶后,通过乙酸乙酯打浆除去大蒜中脂溶性成分,以蒸馏水为提取液,采用响应面法研究料液比、提取温度、提取时间对蒜氨酸提取率的影响。结果表明回归模型能较好的预测各因素与响应值之间的关系,所优化的最佳工艺为:料液比为1∶5,提取温度为32℃,提取时间为70 min。此时蒜氨酸的提取率为92.85±0.63%。     

Effect of thioglycolic acid on Allium sativum L. root meristems]

Thioglycolic acid is, in the Allium test, somewhat less cytotoxic than mercaptoethanol but the characteristics of cytotoxicity are the same in both cases : the mitoclasic effect cannot be separated from a strong mitodepressive effect and is quickly followed by a lethal effect.