Optimization of ultrasonic extraction and clean-up protocol for the determination of polycyclic aromatic hydrocarbons in marine sediments by high-performance liquid chromatography coupled with fluorescence detection

The procedures of ultrasonic extraction and clean-up were optimized for the determination of polycyclic aromatic hydrocarbons (PAHs) in marine sediments. Samples were ultrasonically extracted, and the extracts were purified with a miniaturized silica gel chromatographic column and analyzed with high performance liquid chromatography (HPLC) with a fluorescence detector. Ultrasonication with methanol-dichloromethane (2:1, v/v) mixture gave higher extraction efficiency than that with dichloromethane. Among the three elution solvents used in clean-up step, dichloromethane-hexane (2:3, v/v) mixture was the most satisfactory. Under the optimized conditions, the recoveries in the range of 54.82% to 94.70% with RSDs of 3.02% to 23.22% for a spiked blank, and in the range of 61.20% to 127.08% with RSDs of 7.61% to 26.93% for a spiked matrix, were obtained for the 15 PAHs studied, while the recoveries for a NIST standard reference SRM 1941b were in the range of 50.79% to 83.78% with RSDs of 5.24% to 21.38%. The detection limits were between 0.75 ng L-1 and 10.99 ng L-1for different PAHs. A sample from the Jiaozhou Bay area was examined to test the established methods.     

高效液相色谱法检测水产品中的孔雀石绿和结晶紫

样品中的孔雀石绿、结晶紫经试剂盒提取,浓缩后用经固相萃取柱净化、硼氢化钾还原、反向色谱柱分离,使用荧光检测器检测,孔雀石绿、结晶紫的加标回收率在76.1%～92.5%之间,相对标准偏差1.9%～5.8%,检出限均为0.5μg/kg。结果表明:该方法检测孔雀石绿(MG)、结晶紫(GV)处理简单,灵敏度高,节省时间,可用于大量样品的快速分析。     

Preliminary validation of high performance liquid chromatography method for detection of methyl-testosterone residue in carp muscle

The use of synthetic anabolic steroid methyltestosterone （MT） as growth promoter is prohibited in China. Validations of analytical methods for MT residue in food and the results obtained have become indispensable. The high performance liquid chromatography （HPLC） method for the detection of MT with liquid-liquid extraction by trichloromethanemethanol in carp muscle tissue was preliminarily validated with reference to the following parameters： recovery （accuracy） at the 1, 5 and 10 mgkg^-1 level, between-run and within-run CV values （repeatability, also called relative standard deviation （RSD）） and limit of detection. The recoveries were above 80% and the between-run and within-run CV values below 10% for muscle tissue. The limit of detection was 0.05 mgkg^-1.     

Determination of Steroid Hormone Residues in Fish Tissues Using Gel Permeation Chromatography and Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry

A highly sensitive method was developed for the simultaneous determination of 8 steroid hormones in high-fat fish tissues using ultra high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS).The 8 steroid hormones were extracted from the tissues with diethyl ether.Differing from other common purification methods,the extract solutions were cleaned by gel permeation chromatography(GPC) using ethyl acetate-cyclohexane solution(1:1,v/v) as the mobile phase.The separation of target compounds was carried out by a BEH C18 column and a gradient elution consisting of acetonitrile and 0.2% aqueous formic acid(v/v).The compounds were detected under the multiple reaction monitoring(MRM) mode and quantified with external standard method.This method was validated with respect to linearity,specificity,accuracy and precision.A linearity with correlation coefficient larger than 0.995 was achieved in the range of 0.5 to 50 ng m L~(-1).The average recoveries at the spiked levels of 1.0,5.0,and 10.0 μg kg~(–1) varied between 81.7% and 90.8%,with the relative standard deviations(n=5) ranged from 3.50% to 10.0%.The limit of quantification(LOQ) for 8 steroid hormones ranged from 0.2 to 1.5 μg kg~(-1).It was concluded that this method can be successfully applied for the determination of 8 steroid hormones in complicated matrices including high-fat fish tissues.     

海南岛北部种植园土壤中多环芳烃的质量分数、来源及生态风险

海南岛是我国主要的热带农作物种植基地,对海南岛种植园土壤污染物评估具有重要意义。对海南岛北部种植园土壤中的16种多环芳烃（PAHs）质量分数、来源及生态风险进行评估。使用气相色谱-质谱联用技术对咖啡、香蕉、山药、甘蔗、橙子、百香果和石榴种植园中100件表层土壤样品进行PAHs质量分数测定。结果表明,研究区种植园土壤平均∑₁₆PAHs为37.6 ng/g,低于我国土壤背景值和荷兰无污染土壤限值（< 200 ng/g）。三环、四环和五环PAHs为主要组成成分,分别占据∑₁₆PAHs质量分数的25.8%,26.8%和23.5%。源识别显示,种植园内PAHs主要来自机动车尾气排放,部分来自石油产品的泄露及煤和生物质的低温燃烧。生态风险评价结果显示,等效毒性当量∑₁₆BaP_eq低于国家标准,表明土壤中PAHs污染物水平处于可接受范围。      

邻苯二甲醛柱前衍生高效液相色谱法测定马氏珠母贝中牛磺酸含量

应用邻苯二甲醛柱前衍生高效液相色谱法,测定了马氏珠母贝中牛磺酸,结果表明:采用邻苯二甲醛为柱前衍生剂,与原料中的牛磺酸发生衍生化反应后,以甲醇-乙酸钠溶液(体积比40︰60)为流动相,使用ODSC18分离柱,FLD检测器(EX=338nm,EM=444nm)等色谱条件下检测,牛磺酸的线性范围为2.0～14.0μg/mL,相关系数R2为0.9984,回收率为97.9%～103.0%,RSD=2.146%(n=6),检出限为0.77μg(S/N=3)。     

Concentrations and sources of polycyclic aromatic hydrocarbons in topsoil of Benxi City,Northeast China

Polycyclic aromatic hydrocarbons (PAHs) contamination of topsoil in Benxi City, Northeast China was investigated in this study, and sampling sites were located in industrial area and residential area. Results demonstrate that there is high variability in the total PAHs concentrations, ranging from 783.00 ng/g to 729 076.29 ng/g dry weight in the topsoil of Benxi City and the pollution in industrial area is the most serious. The results also present that higher molecular weight PAHs (4–6 rings) contribute substantially (79.03%) to the overall content of PAHs. The PAHs sources were determined with factor analysis by nonnegative constraints, and the results show that PAHs originating from traffic tunnel, power plant, coke oven and residential emission sources, account for 27.10%, 40.81%, 20.11% and 11.98%, respectively, of the total. The PAHs pollution is the most serious around Benxi Iron and Steel Group Corporation, and the PAHs mainly originate from coke oven and traffic tunnel, with the average contribution rate of 57.40% and 42.60%, respectively. The source apportionment results are basically consistent with the industry distribution, and the geographical and climatic characteristics of the study area.     

Molecularly imprinted polymers for selective extraction of crystal violet from natural seawater coupled with high-performance liquid chromatographic determination

Molecularly imprinted polymers(MIPs) were prepared by the bulk polymerization using crystal violet as the template molecule, and the methacrylic acid and ethylene glycol dimetheacrylate as functional monomer and cross-linker, respectively. Systematic investigations of synthetic conditions were conducted. The surface morphology and recognition mechanism of the obtained polymers were studied using scanning electron microscope and spectrophotometric analysis. MIPs showed high affinity to template molecule and were successfully applied as special solid-phase extraction sorbent for selective extraction of crystal violet from natural seawater. An off-line molecularly imprinted solid-phase extraction(MISPE) method followed by high-performance liquid chromatography with diodearray detection for the analysis of crystal violet was also established. MISPE columns have good recoveries for crystal violet standard solutions and good linearity was obtained over the concentration range of 0-200 μg L-1(R2 0.99). Finally, two natural seawater samples were investigated. The recoveries of spiked seawater on the MISPE columns were from 44.47% to 62.34%, the relative standard deviation(n=3) being in the range of 2.89%-5.96%.     

Simultaneous identification and quantification of tetrodotoxin in fresh pufferfish and pufferfish-based products using immunoaffinity columns and liquid chromatography/quadrupole-linear ion trap mass spectrometry

In this study,we established a comprehensive method for simultaneous identification and quantification of tetrodotoxin(TTX) in fresh pufferfish tissues and pufferfish-based products using liquid chromatography/quadrupole-linear ion trap mass spectrometry(LC-QqLIT-MS).TTX was extracted by 1% acetic acid-methanol,and most of the lipids were then removed by freezing lipid precipitation,followed by purification and concentration using irnmunoaffinity columns(IACs).Matrix effects were substantially reduced due to the high specificity of the IACs,and thus,background interference was avoided.Quantitation analysis was therefore performed using an external calibration curve with standards prepared in mobile phase.The method was evaluated by fortifying samples at 1,10,and 100 ng/g,respectively,and the recoveries ranged from 75.8%-107%,with a relative standard deviation of less than 15%.The TTX calibration curves were linear over the range of 1-1 000 ng/L,with a detection limit of 0.3 ng/g and a quantification limit of 1 ng/g.Using this method,samples can be further analyzed using an informationdependent acquisition(IDA) experiment,in the positive mode,from a single liquid chromatography-tandem mass spectrometry injection,which can provide an extra level of confirmation by matching the full product ion spectra acquired for a standard sample with those from an enhanced product ion(EPI) library.The scheduled multiple reaction monitoring method enabled TTX to be screened for,and TTX was positively identified using the IDA and EPI spectra.This method was successfully applied to analyze a total of 206 samples of fresh pufferfish tissues and pufferfish-based products.The results from this study show that the proposed method can be used to quantify and identify TTX in a single run with excellent sensitivity and reproducibility,and is suitable for the analysis of complex matrix pufferfish samples.     

超声波辅助提取河豚毒素的研究

以月腹刺鲀(Gastrophysus lunaris)卵巢为原料,利用超声波辅助提取河豚毒素(Tetrodotoxin,TTX)。通过单因素实验及二次回归旋转组合设计实验研究液料比、超声温度、超声时间等因素对提取率的影响,确定超声波辅助提取河豚毒素的最佳工艺条件。结果表明:在实验范围内影响河豚毒素得率的各因素大小依次为液料体积质量比、超声温度、超声时间;超声波辅助提取河豚毒素的最佳工艺条件为液料体积(mL)质量(g)比3.03∶1,超声温度39℃,超声时间22 min,该工艺条件下提取河豚毒素得率为63.57μg/g。     

Isolation,Identification, and Quantitative Determination of Saponin in Apostichopus japonicus by HPLC-DAD

Apostichopus japonicus is an important invertebrate that is widely used as a tonic food in Asian countries.The purpose of this study is to purify and identify a class of compound,the saponins,from the body wall of A.japonicus,and to establish a new me-thod to determine the quantity of saponins in the sea cucumber.In this study,the saponins of A.japonicus,cladoloside A(CA),were ob-tained from 80%ethanol extract by column chromatography for the first time and were characterized using the spectral method.The resulting purified saponins were then profiled using ¹HNMR,¹³CNMR,and ESI-MS,which revealed the CA molecular formula of C₅₃H₈₂O₂ and contained a triterpenoid backbone,a methylglucopyranosyl moiety,a quinovopyranosyl,and two xylopyranosyls.A me-thod for the quantitative determination of CA,comprising microwave-assisted extraction,high-performance liquid chromatography,and diode array detector method,was established.Extraction efficiency was optimized by changing microwave power,extraction sol-vent,volume,time,and temperature.Results showed that under the optimum conditions(extraction time of 10 min,temperature of 45℃,and solvent of 25 mL 70%ethanol under 400 W),the detection limit of CA was 0.0015 mg mL^?1,and the recoveries of CA from samples at spiking levels of 10,20,and 50μg g^?1 ranged from 90.1%-104%.The proposed method was successfully applied to ana-lyze the saponins in different tissues of A.japonicus collected in different seasons.The method developed in this study can provide quantitative technical support for the quality control of A.japonicus.     

Determination of lignin in marine sediment using alkaline cupric oxide oxidation-solid phase extraction-on-column derivatization-gas chromatography

Lignin serves as one of the most important molecular fossils for tracing Terrestrial Organic Matters (TOMs) in marine environment. Extraction and derivatization of lignin oxidation products (LOPs) are crucial for accurate quantification of lignin in marine sediment. Here we report a modification of the conventional alkaline cupric oxide (CuO) oxidation method, the modification consisting in a solid phase extraction (SPE) and a novel on-column derivatization being employed for better efficiency and reproducibility. In spiking blanks, recoveries with SPE for the LOPs are between 77.84% and 99.57% with relative standard deviations (RSDs) ranging from 0.57% to 8.04% (n=3), while those with traditional liquid-liquid extraction (LLE) are from 44.52% to 86.16% with RSDs being from 0.53% to 13.14% (n=3). Moreover, the reproducibility is greatly improved with SPE, with less solvent consumption and shorter processing time. The average efficiency of on-column derivatization for LOPs is 100.8% ± 0.68%, which is significantly higher than those of in-vial or in-syringe derivatization, thus resulting in still less consumption of derivatizing reagents. Lignin in the surface sediments sampled from the south of Yangtze River estuary, China, was determined with the established method. Recoveries of 72.66% to 85.99% with standard deviation less than 0.01mg/10g dry weight are obtained except for p-hydroxyben-zaldehyde. The lignin content Σ8 (produced from 10 g dry sediment) in the research area is between 0.231 and 0.587 mg. S/V and C/V ratios (1.028 ± 0.433 and 0.192 ± 0.066, respectively) indicate that the TOMs in this region are originated from a mixture of woody and nonwoody angiosperm plants; the high values of (Ad/Al) v suggest that the TOMs has been highly degraded.     

Screening of Extraction Methods for Glycoproteins from Jellyfish （Rhopilema esculentum） Oral-Arms by High Performance Liquid Chromatography

In order to select an optimum extraction method for the target glycoprotein (TGP) from jellyfish (Rhopilema esculentum) oralarms, a high performance liquid chromatography (HPLC)–assay for the determination of the TGP was developed. Purified target glycoprotein was taken as a standard glycoprotein. The results showed that the calibration curves for peak area plotted against concentration for TGP were linear (r = 0.9984, y = 4.5895x+47.601) over concentrations ranging from 50 to 400 mgL-1. The mean extraction...     

Extraction of organochlorine pesticides in sediments using soxhlet,ultrasonic and accelerated solvent extraction techniques

The application of soxhlet, ultrasonic and accelerated solvent extraction techniques to the analysis of six organochlorine pesticides (α-HCH, β-HCH, γ-HCH, o, p'-DDT, p,p'-DDT and p, p'-DDE) in Taihu Lake sediment samples is described. It was found that the limits of quantification ranged from 0.002μgg-1 to 0.004μgg-1, and the recoveries of organochlorine pesticides with the three extraction techniques were acceptable (>80.7%). With a mass selective detector, better results were obtained by accelerated solvent extraction using hexane-acetone (1:1) as compared with soxhlet and ultrasonic extraction. It was shown that the accelerated solvent extraction was the optimum technique for the analysis of organochlorine pesticides in sediments. The general features of the three extraction techniques are also presented.     

Simultaneous determination of iron and manganese in water using artificial neural network catalytic spectrophotometric method

A new analytical method using Back-Propagation (BP) artificial neural network and kinetic spectrophotometry for simultaneous determination of iron and magnesium in tap water, the Yellow River water and seawater is established. By conditional experiments, the optimum analytical conditions and parameters are obtained. Levenberg-Marquart (L-M) algorithm is used for calculation in BP neural network. The topological structure of three-layer BP ANN network architecture is chosen as 15-16-2 (nodes). The initial value of gradient coefficient μ is fixed at 0.001 and the increase factor and reduction factor of μ take the default values of the system. The data are processed by computers with our own programs written in MATLAB 7.0. The relative standard deviation of the calculated results for iron and manganese is 2.30% and 2.67% respectively. The results of standard addition method show that for the tap water, the recoveries of iron and manganese are in the ranges of 98.0%-104.3% and 96.5%-104.5%, and the RSD is in the range of 0.23%-0.98%; for the Yellow River water (Lijin district of Shandong Province), the recoveries of iron and manganese are in the ranges of 96.0%-101.0% and 98.7%-104.2%, and the RSD is in the range of 0.13%-2.52%; for the seawater in Qingdao offshore, the recoveries of iron and manganese are in the ranges of 95.3%-104.8% and 95.3%-104.7%, and the RSD is in the range of 0.14%-2.66%. It is found that 21 common cations and anions do not interfere with the determination of iron and manganese under the optimum experimental conditions. This method exhibits good reproducibility and high accuracy in the determination of iron and manganese and can be used for the simultaneous determination of iron and manganese in tap water and natural water. By using the established ANN- catalytic spectrophotometric method, the iron and manganese concentrations of the surface seawater at 11 sites in Qingdao offshore are determined and the level distribution maps of iron and manganese are drawn.     

Screening of extraction methods for glycoproteins from jellyfish (Rhopilema esculentum) oral-arms by high performance liquid chromatography

In order to select an optimum extraction method for the target glycoprotein (TGP) from jellyfish (Rhopilema esculentum) oral-arms, a high performance liquid chromatography (HPLC)-assay for the determination of the TGP was developed. Purified target glycoprotein was taken as a standard glycoprotein. The results showed that the calibration curves for peak area plotted against concentration for TGP were linear (r = 0.9984, y = 4.5895x+47.601) over concentrations ranging from 50 to 400 mgL-1. The mean extraction recovery was 97.84% (CV2.60%). The fractions containing TGP were isolated from jellyfish (R. esculentum) oral-arms by four extraction methods: 1) water extraction (WE), 2) phosphate buffer solution (PBS) extraction (PE), 3) ultrasound-assisted water extraction (UA-WE), 4) ultrasound-assisted PBS extraction (UA-PE). The lyophilized extract was dissolved in Milli-Q water and analyzed directly on a short TSK-GEL G4000PWXL (7.8 mm×300 mm) column. Our results indicated that the UA-PE method was the optimum extraction method selected by HPLC.     

ICP-AES测定金属元素铜活动态及其初步应用

金属元素活动态是我国5种深穿透地球化学技术中应用最广的方法之一。此方法是通过提取地表土壤一定深度介质中元素活动态的微弱隐伏矿信息而找寻隐伏矿,属于通过直接信息找矿,目前该方法的标准化程度不高。为将铜元素活动态应用于地质找矿,该文对铜元素4个活动态的相态(水浸出态、粘土吸附态、有机链合态、铁锰氧化物态)循序提取的条件及ICP-AES测定技术进行了研究,包括采样深度、分析样品粒度、样品存放时间、提取条件、测定干扰等,通过对铜活动态提取液处理方法的改进,提高了分析速度和分析数据的精度。由实验数据计算出各相态方法的检出限和精密度RSD(n=10),水浸出态为0.079mg/L,13.90%~22.49%;粘土吸附态为0.275mg/L,11.99%~23.29%;有机链合态为0.362mg/L,10.70%~57.27%;铁锰氧化物态为0.230mg/L,17.83%~38.61%。该方法操作方便,精密度较好,应用于碑楼隐伏铜矿的野外探测实验,探测结果所圈定的综合异常与实际矿体的水平投影相符,取得了较满意的试验效果,该方法可应用于不同地质景观区。     

Distribution of organochlorine pesticides and polychlorinated biphenyls in Ny-Alesund of the Arctic

Sediment, moss and animal-dropping samples were collected from pesticides (OCPs) and polychlorinated biphenyls (PCBs) in the Arctic area. OCPs and PCBs were analyzed using the capillary gas chromatography (GC-ECD). Recoveries of three surrogates were at a range of 69.5～92.2% (mean: 78.8%), 75.1～112.1% (98.5%) and 67.7～95.1% (79.5%), and the relative standard deviations were 6.7%, 8.0% and 11.3%, respectively. The major monomers of OCPs were hexachlorocyclohexanes (HCHs) and DDTs. Their concentrations were at a range of 0.86～4.50 ng/g (mean: 2.24 ng/g) and 0.22～1.09 ng/g (mean: 0.55 ng/g). The concentrations of PCBs were 0.20～3.25 ng/g (mean: 0.84 ng/g) and the major congeners were tetra-, penta-, and hexa-PCBs, which were up to 23.8%、26.7% and 32.3% of total PCBs, respectively. Based on the results of the ratio analysis and the principal component analysis, the atmospheric transport is believed be one     

Preparation of polysaccharides in different molecular weights from Ulva pertusa Kjellm (Chorophyta)

As molecular weight controls the biological activities of polysaccharides, screening the optimal molecular weight of polysaccharides is important in drug research and application. In this study, hydrogen peroxide was employed as oxidant, and temperature, reaction time, and concentration of polysaccharides and hydrogen peroxide were examined for their effects on the preparation of polysaccharides in different molecular weights fromUlva pertusa. Our experiment suggested that the optimal degradation concentrations for polysaccharides and hydrogen peroxide were 2.5% (w/v) and 8.0% (v/v), respectively. The range of degradation measured by relative viscosity was mainly controlled by temperature and time. Results revealed that 35°C was the optimal temperature for obtaining low-degradation samples, and 50°C was the most favorable temperature to accelerate the reaction to yield highly-degraded samples. Four samples in different molecular weights A, B, C and D were finally prepared. The controllability was evaluated by the relative standard deviation (RSD) of relative viscosity, and the peak molecular weights and the polydispersity indexes (M_w/M_n) of molecular weights were measured by high performance gel permeation chromatography (HPGPC). This work was supported by the Scientific and Technical Bureau of Shandong Province.     

Studies on the antitumor activity and isolation of the glycoprotein from the gonad ofChlamys (Azumapecten) farreri

After being degreased with acetone, the female gonad ofChlamys (Azumapecten) farreri was extracted with water, and then subjected to DEAE—cellulose column chromatography and purified by Sephadex G-100 chromatography to obtain a pure female gonad glycoprotein (FGGP). The IR spectra, electrophoresis and chemical analysis of FGGP indicated that it was a kind of acid glycoprotein and that the content of total protein and sugar were 54.8% and 40%, respectively. Pharmacological tests showed that FGGP could inhibit (inhibition rate of 40.87%) the growth of Sarcoma 180 of mice.