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Ciguatoxins (CTXs), produced by toxic benthic dinoflagellates, can bioaccumulate in marine organisms at higher trophic levels. The current study evaluated the uptake and depuration kinetics of some of the most potent CTXs, Pacific CTX-1, -2, and -3 (P-CTX-1, -2, and -3), in orange-spotted grouper (Epinephelus coioides) exposed to 1 ng P-CTXs g-1 fish daily. Over a 30 d exposure, P-CTX-1, -2, and -3 were consistently detected in various tissues of exposed fish, and the concentrations of the total P-CTXs in tissues generally ranked following the order of liver, intestine, gill, skin, brain, and muscle. Relatively higher uptake rates of P-CTX-1 in the groupers were observed compared with those of P-CTX-2 and -3. The depuration rate constants of P-CTX-1, -2, and -3 in different tissues were (0.996-16.5) × 10-2, (1.51-16.1) × 10-2, and (0.557-10.6) × 10-2 d-1, respectively. The accumulation efficiencies of P-CTX-1, -2, and -3 in whole groupers were 6.13%, 2.61%, and 1.15%, respectively. The increasing proportion of P-CTX-1 and the decreasing proportion of P-CTX-2 and -3 over the exposure phase suggest a likely biotransformation of P-CTX-2 and -3 to P-CTX-1, leading to higher levels of P-CTX-1 in fish and possibly a higher risk of CTXs in long-term exposed fish.

Because only very weak signals of fragment ions of nosiheptide can be obtained, nosiheptide is usually detected by liquid chromatography-tandem mass spectrometry (LC-MS/MS) via the determination of its hydrolyzed degradation product named HMIA in previous studies. The indirect method suffers from several problems, such as complicated samplepreparation, unavailable commercial HMIA, and the risk of the false-positive result by HMIA. However, we found that nosiheptide could produce several significant fragment ions under high collision energy (CE). Therefore, we developed a method for the direct determination of nosiheptide by LC-MS/MS in animal tissues. The sample was extracted with ACN, then degreased with n-hexane, and purified by an HLB solid-phase extraction (SPE) cartridge. After being filtered through the PTFE filter, it was analyzed by LC-MS/MS in selected reaction monitoring (SRM) mode. The influencing factors, such as mobile phase, SPE cartridge, filter material, and matrix effect, were investigated. Nosiheptide showed a good linear relationship (R

The occurrence, spatial distribution, and partitioning behavior of 17 marine lipophilic phycotoxins (MLPs) in surface and bottom seawater, particulate organic matter (POM), and surface sediment from the Pearl River Estuary (PRE) were investigated to understand current contamination and the potential risks to marine ecosystems in this region. Nine MLPs were detected, including azaspiracid1-3, gymnodimine, okadaic acid, dinophysistoxin 1-2, pectenotoxin2 (PTX2), and homoyessotoxin, with Σ

The chromophores responsible for light absorption in atmospheric brown carbon (BrC) are not well characterized, which hinders our understanding of BrC chemistry, the links with optical properties, and accurate model representations of BrC to global climate and atmospheric oxidative capacity. In this study, the light absorption properties and chromophore composition of three BrC fractions of different polarities were characterized for urban aerosol collected in Xi'an and Beijing in winter 2013-2014. These three BrC fractions show large differences in light absorption and chromophore composition, but the chromophores responsible for light absorption are similar in Xi'an and Beijing. Water-insoluble BrC (WI-BrC) fraction dominates the total BrC absorption at 365 nm in both Xi'an (51 ± 5%) and Beijing (62 ± 13%), followed by a humic-like fraction (HULIS-BrC) and high-polarity water-soluble BrC. The major chromophores identified in HULIS-BrC are nitrophenols and carbonyl oxygenated polycyclic aromatic hydrocarbons (OPAHs) with 2-3 aromatic rings (in total 18 species), accounting for 10% and 14% of the light absorption of HULIS-BrC at 365 nm in Xi'an and Beijing, respectively. In comparison, the major chromophores identified in WI-BrC are PAHs and OPAHs with 4-6 aromatic rings (in total 16 species), contributing 6% and 8% of the light absorption of WI-BrC at 365 nm in Xi'an and Beijing, respectively.

Ciguatoxins (CTXs), produced by toxic benthic dinoflagellates, can bioaccumulate in marine organisms at higher trophic levels. The current study evaluated the uptake and depuration kinetics of some of the most potent CTXs, Pacific CTX-1, -2, and -3 (P-CTX-1, -2, and -3), in orange-spotted grouper (

Introduction Crotalaria sessiliflora L. is a Chinese traditional herb for treatment of cutaneum carcinoma and cervical carcinoma. In addition to monocrotaline, coexisting pyrrolizidine alkaloids (PAs) also require further quantification for quality control and pharmaceutical uses of the herb. Objective To establish a UPLC-Q-Orbitrap/MS method of simultaneous determination of coexisting PAs with same parent structure for quality control and comprehensive researches of Crotalaria sessiliflora L. Methodology PAs in Crotalaria sessiliflora L. were analysed by UPLC-Q-Orbitrap/MS method. Coexisting PAs were identified by mass data of full MS-dd-MS2 based on the characteristic fragmentation pattern and necine-core structure. Moreover, quantification of PAs was conducted by parallel reaction monitoring (PRM) mode using m/z 138, m/z 120 and m/z 94 from identical necine-core structure as quantitative ions with single monocrotaline standard for accurate calibration. Results Five PAs, named monocrotaline, retrorsine, senecionine, integerrimine, O-9-angeloylretronecine, were indentified and confirmed. Quantitative ions of m/z 138, m/z 120 and m/z 94 were used for quantification of PAs containing the necine-core structure in Crotalaria sessiliflora L. The results demonstrated that contents, precision and recoveries of the five PAs mentioned earlier were respectively 3.307–30.35 μg/g, 1.1–4.5% and 88.91–92.33% while using m/z 120 as the best quantitative ion. Conclusion The UPLC-Q-Orbitrap/MS method was established for simultaneous determination of five PAs in Crotalaria sessiliflora L. without all corresponding standards, and was proved that it was simple, convenient and effective for comprehensive quality control and pharmaceutical uses. Copyright © 2017 John Wiley & Sons, Ltd.

The sources, formation mechanism and amount of organosulfates (OS) in atmospheric aerosol are not yet well understood, partly due to the lack of authentic standards for quantification. In this study, we report an improved robust procedure for the synthesis of organosulfates with different functional groups. Nine authentic organosulfate standards were synthesized and four standards (benzyl sulfate, phenyl sulfate, glycolic acid sulfate, and hydroxyacetone sulfate) were used to quantify their ambient concentrations. The authentic standards and ambient aerosol samples were analyzed using an optimized ultra performance liquid chromatography–electrospray ionization-tandem mass spectrometric method (UPLC–ESI–MS/MS). The recovery ranged from 80.4 to 93.2 %, the limits of detection and limits of quantification obtained were 1.1–16.7 and 3.4–55.6 pg m−3, respectively. Measurements of ambient aerosol particle samples collected in winter 2013/2014 in urban Xi'an, northwestern China, show that glycolic acid sulfate (77.3 ± 49.2 ng m−3) is the most abundant species of the identified organosulfates followed by hydroxyacetone sulfate (1.3 ± 0.5 ng m−3), phenyl sulfate (0.14 ± 0.09 ng m−3), and benzyl sulfate (0.04 ± 0.01 ng m−3). Except for hydroxyacetone sulfate, which seems to form mainly from biogenic emissions in this region, the organosulfates quantified during winter in Xi'an show an increasing trend with an increase in the mass concentrations of organic carbon indicating their anthropogenic origin.

The sources, formation mechanism and amount of organosulfates (OS) in atmospheric aerosol are not yet well understood, partly due to the lack of authentic standards for quantification. In this study, we report an improved robust procedure for the synthesis of organosulfates with different functional groups. Nine authentic organosulfate standards were synthesized and four standards (benzyl sulfate, phenyl sulfate, glycolic acid sulfate, and hydroxyacetone sulfate) were used to quantify their ambient concentrations. The authentic standards and ambient aerosol samples were analyzed using an optimized ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometric method (UPLC–ESI–MS/MS). The recovery ranged from 80.4 % to 93.2 %, the limits of detection and limits of quantification obtained were 1.1–16.7 pg m−3 and 3.4–55.6 pg m−3, respectively. Measurements of ambient aerosol particle samples collected in winter 2013–2014 in urban Xi’an, northwest China, show that glycolic acid sulfate (77.3 ± 49.2 ng m−3) is the most abundant species of the identified organosulfates followed by hydroxyacetone sulfate (1.3 ± 0.5 ng m−3), phenyl sulfate (0.14 ± 0.09 ng m−3), and benzyl sulfate (0.04 ± 0.01 ng m−3). Except for hydroxyacetone sulfate, which seems to form mainly from biogenic emissions in this region, the organosulfates quantified during winter in Xi’an show an increasing trend with an increase in the mass concentrations of organic carbon indicating their anthropogenic origin.

An integrated low-cost alloy and corrosive anion system was attempted to perform aqueous hydrodechlorination (HDC) of biorefractory chlorinated organics. Several influencing factors were investigated for HDC of 2-chlorophenol (2-CP) including anion species, alloy categories, and their dosages. Al–Ni alloy presented the remarkably highest reactivity to remove 2-CP under the action of aqueous fluorine ion (F − ) compared with other anions and alloys used in this study. Increasing Al–Ni alloy dosage or initial concentration of F − could enhance of 2-CP HDC efficiency. The optimized results showed that 0.389 mM of 2-CP in solution could be completely dechlorinated into phenol using 10 g L −1 of Al–Ni alloy and 39.7 mM initial concentration of F − within 120-min reaction. The used Al–Ni alloy after each HDC reaction could almost be reactivated to the original activity level of raw Al–Ni alloy for HDC after Ca(OH) 2 treatment to remove the adsorbed species such as AlF 3 on alloy surface during three cycle reaction. Moreover, after Ca(OH) 2 activation, F − in the effluent solution could also be effectively removed to under 0.8 mg L −1 due to the formation of insoluble CaF 2 . The maintenance and regeneration of Al–Ni alloy activity for HDC was mainly attributed to the effects of Ca(OH) 2 activation to remove adsorbed species on Al–Ni surface, inherent structure of alloy with Ni droplets homogeneously dispersed into Al substrate, and the change of its surface morphology with more active sites produced by F − corrosion.

A method was developed for the determination of 10 androgens, 11 progesterones, 10 glucocorticoids, 5 estrogens and 5 resorcylic acid lactones in cereal feeds by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Ethyl acetate was used to extract the sample, and the mixed QuEChERS sorbents were dispersed for purification. The extract was separated by a C18 column and detected in positive or negative multiple reaction monitoring (MRM) mode. With the optimal conditions, the linear ranges of 41 hormones were 5.0 - 200.0 microg/kg with the correlation coefficients above 0.99 and the limits of quantification (LOQs, S/N > or = 10) were 0.123- 2.72 microg/kg. The method validation was carried out at three spiking levels, and the recoveries were 70.3% - 118.1% for soybean meals with the relative standard deviations (RSDs) of 0.82% - 19.5%, and 76.1% - 122.8% for corn meals with the RSD of 1.3% - 15.0%. This method is simple, fast and credible, can be applied to simultaneous screening and determination of estrogens, progesterones, androgens, glucocorticoids and resorcylic acid lactones belonging to the illegal additive of hormones in cereal feeds.

It has recently been suggested that acetylcholine plays an important role in the modulation of tissue inflammation. In order to further understand the newly discovered cholinergic anti-inflammatory pathway, tracking the concentration changes of acetylcholine in tissue is required. This paper describes the development of a method coupling hydrophilic interaction chromatography with electrospray ionization tandem mass spectrometry (HILIC/ESI-MS/MS) for the separation and quantification of acetylcholine in microdialysis samples of normal rats and of rats with local inflammation. The separation of acetylcholine from interferential endogenous compounds and inorganic cations was achieved with a zwitterionic stationary phase column using isocratic elution. Low-energy collision-induced dissociation tandem mass spectrometric (CID-MS/MS) analysis was carried out in the positive ion mode using multiple reaction monitoring (MRM) of the following mass transitions: m/z 146 --> 87 for acetylcholine and m/z 155 --> 87 for the internal standard acetylcholine-D9. The limit of detection for acetylcholine was found to be 0.075 fmol on-column with a signal-to-noise ratio of 3:1. The lower limit of quantification was 0.25 fmol on-column. The calibration curves obtained for acetylcholine in blank microdialysates were linear in the ranges of 0.025-50 nM and 0.025-0.5 nM, with correlation coefficients equal to or greater than 0.9994 and 0.9969, respectively. The recoveries of acetylcholine for high (2 nM) and low (0.5 nM) concentrations were in the ranges of 90-96% and 95-109%, respectively. The coefficients of variation for intra-day and inter-day reproducibility were equal to or less than 7.3% and 10.4%, respectively. The method has been successfully applied in the measurement of acetylcholine in microdialysates from normal and inflamed rat tissue.

Although positive-ion (PI) electrospray ionisation mass spectrometry (ESI-MS) has been usually applied for the analysis of native metallothioneins (MT) isoforms binding with metal ions, it suffers from the lack of insensitivity under neural conditions because of the low efficiency of protonation during ESI process. In this study, multiply deprotonated metallothionein (Zn7-MT-2a), produced from tris(hydroxymethyl)aminomethane (TRIS) acetate solutions under near neutral condition, was analyzed by negative-ion (NI) ESI-MS. Compared with the ammonium acetate buffer system which has been normally used for ESI-MS experiments, the use of TRIS acetate buffer results in the formation of more abundant ions and higher charge states of MT-2a, and consequently higher intensity is attained. The sensitivity enhancement of the system could be explained by the high gas-phase proton affinity, small molecular volume of the anion (acetate), and the high hydrogen consumption by TRIS in the buffer system, which would all favor the deprotonation of the analyte during NI-ESI process. Factors that may affect the NI-ESI responses, such as acid added in buffer, pH and concentration of TRIS acetate had been evaluated, and the result showed that using acetic acid, lower pH under the pH range evaluated, higher concentration of TRIS acetate all favored the MS detection of MT-2a. Our finding sheds light on a buffer system that may offer substantial sensitivity advantages in the studying of weakly bound, non-covalent complexes such as metal binding MTs, which are usually analyzed under near neutral conditions.

Mercury, a highly toxic metal found widely throughout the environment, is a potent inducer of metallothionein (MT) expression. The role of MTs in the detoxification of mercury after its oral intake in mammals is studied. After feeding rats with mercuric chloride by gastric gavage, the distribution of heavy metals in rat tissues was investigated by inductively coupled plasma mass spectrometry (ICP-MS). Extensive accumulation of mercury, copper and zinc in kidney and liver is observed. A homemade preparative size-exclusion chromatography (SEC) column (30 cm × 1.9 cm) packed with Sephadex G-75 (40–120 µm particle size) gel (Pharmacia) was used for the purification of MT fractions in rat tissues. Preliminary results from SEC indicate that the mercury-binding MT levels in liver were much lower than in kidney. The MT fractions were collected, desalted, and then separated by reversed-phase high-performance liquid chromatography (HPLC) with UV–Vis spectrometry, ICP-MS and electrospray ionization MS detection. One major and several minor peaks were observed in the HPLC chromatograms of the MT fraction for the kidney sample. UV absorption spectra indicate that MTs were found to bind with mercury. There were no significant mercury-binding MTs detected in the liver sample using UV detection. ICP-MS detection showed that mercury-binding MTs in kidney contained large amounts of mercury and copper but little zinc. Further characterization with ESI-MS showed that the major peak found in kidney contained Hg6Cu, Hg5Cu2-MT-2c and Hg6-MT-2β, Hg6Cu-MT-1γ, Hg7-MT-2α. However, distinction between copper and zinc could not be made based on current mass spectrometric analysis because of instrumental resolution limitations. Copyright © 2005 John Wiley & Sons, Ltd.

The multi-metal-binding MT fractions in rat tissues after oral intake of cinnabar were characterized by hyphenated size-exclusion chromatography (SEC) and inductively coupled plasma mass spectrometry (ICP-MS). With the increase of both the feeding time and dosage of cinnabar the amounts of Hg-binding MT fractions in rat kidney of groups fed cinnabar increased significantly compared with the control group. Meanwhile, more Cu-binding MT were synthesized in the rat kidney, which confirmed the manipulating effect of MT in the homeostasis of Cu for detoxification of nephritic mercury. Although the Hg-binding MT fractions in rat liver of all cinnabar groups were almost independent of cinnabar dosage (2.5-5.0 g kg(-1) bw) and feeding time (2-4 weeks), the amount was much higher than that of the control group. The amounts of Hg-MT in other rat tissues of the cinnabar groups and the control group were compared and their significance is discussed.

Cinnabar, as one of the most widely used mineral drugs in traditional Chinese medicines, has been proven to have prominent curative effects in clinical use for more than 2000 years. But the safety and toxicity of the drug has been under constant debate in clinic usage. Metallothionein (MT) contains about 30% of cysteine in the molecule, and plays an important detoxification role against heavy metals. In this study, it was used as a biomarker to assess mercurial accumulation in rats fed orally with cinnabar. After feeding rats with cinnabar by gastric gavage at different dosages and at different times, the distribution of heavy metals (including mercury, copper and zinc) and MT was investigated among rat tissues, including liver, kidney, heart, brain, testis and blood. Metals and MT determinations were carried out using inductively coupled plasma mass spectrometry (ICP-MS) and a modified mercury saturation assay technique respectively. The results indicated that mercury was easily accumulated in the tissues of rats exposed to cinnabar, especially in kidney. For example: at a feeding dosage of 5 g kg−1 (bw) for 4 weeks, the mercury concentrations in kidney were 13, 8.7, 21.6 and 26 times those in liver, testis, brain and heart respectively; and at 2.5 g kg−1 (bw) for 2 weeks, the mercury concentrations in kidney were 21, 2.1, 3 and 21 times those in liver, testis, brain and heart respectively. In addition, mercury in kidney and liver of all cinnabar groups was significantly higher than that of the control group (P < 0.01). A high positive correlation observed between MT concentrations and mercury levels in both liver and kidney (R2 = 0.9299, P < 0.02 for liver; R2 = 0.9923, P < 0.0008 for kidney) indicated that MT could be used as a biomarker for mercury in tissues. Copyright © 2004 John Wiley & Sons, Ltd.

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the determination of estrone, 17beta-estradiol, estriol in animal liver and kidney tissues. The sample was extracted by tert-butyl methyl ether, and the extract was evaporated by nitrogen at 45 degrees C. The residue was redissolved in hexane-dichloromethane (6:4, v/v), then purified on a silica solid-phase extraction column. The eluant was evaporated by nitrogen, dissolved in acetonitrile-water (7:3, v/v) and then analyzed by LC-MS/MS. The separation of estrogens was performed on a Poroshell 120 EC-C18 column with the mobile phases of acetonitrile and water with a gradient elution. The identification and quantification of estrogens were carried out by negative electrospray ionization in the multiple reaction monitoring mode using external standard method. The calibration curves showed good linearity in the range of 1.0-20.0 microg/kg with the correlation coefficients above 0.99. The limit of quantification was 1.0 microg/kg for each estrogen. The average recoveries of the estrogens spiked at 1.0, 2.0, 10.0 microg/kg levels in different matrices were between 70.2% and 114%, and the relative standard deviations were between 2.01% and 14.5%. The method is simple, rapid, sensitive, good in purification effect. It is suitable for the confirmation and quantification of estrogens in animal liver and kidney tissues.

A method was developed for the determination of 10 androgens, 11 progesterones, 10 glucocorticoids, 5 estrogens and 5 resorcylic acid lactones in cereal feeds by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Ethyl acetate was used to extract the sample, and the mixed QuEChERS sorbents were dispersed for purification. The extract was separated by a C18 column and detected in positive or negative multiple reaction monitoring (MRM) mode. With the optimal conditions, the linear ranges of 41 hormones were 5.0 - 200.0 microg/kg with the correlation coefficients above 0.99 and the limits of quantification (LOQs, S/Nor = 10) were 0.123- 2.72 microg/kg. The method validation was carried out at three spiking levels, and the recoveries were 70.3% - 118.1% for soybean meals with the relative standard deviations (RSDs) of 0.82% - 19.5%, and 76.1% - 122.8% for corn meals with the RSD of 1.3% - 15.0%. This method is simple, fast and credible, can be applied to simultaneous screening and determination of estrogens, progesterones, androgens, glucocorticoids and resorcylic acid lactones belonging to the illegal additive of hormones in cereal feeds.

A method for the determination of metabolite residues of dipyrone, 4-formylaminoantipyrine (FAA), 4-aminoantipyrine (AA) and 4-methylaminoantipyrine (MAA) in bovine muscle and pork muscle has been developed. Homogenized muscle sample was extracted with Na2SO4-Na2SO3 solution. After filtration, the extract was cleaned-up by a C18 solid phase extraction cartridge. Prepared sample was determined using high performance liquid chromatography under the following conditions: Inertsil ODS-3 column, methanol and water as mobile phase with gradient elution, detection wavelength of 265 nm. FAA was quantified by external standard method, while AA and MAA were quantified by internal standard method using isopropylaminoantipyrine as internal standard. Limits of detection of FAA was 12.5 microg/L, while those of AA and MAA were 15.0 microg/L and 20.0 microg/L, respectively. The limit of detection of this method was 50 microg/L for all the three metabolites. The recoveries of FAA, AA and MAA were 81.3% - 92.5%, 82.0% - 96.0%, and 80.4% - 90.6%, respectively, at the spiked levels of 50 - 400 microg/kg. The relative standard deviations of the method were less than 7%.

Polymerization of metallothioneins (MTs) is one of the commonly encountered puzzles in researching the structure and function of metallothioneins. In this work, a method involving SEC coupled with negative ion electrospray ionization mass spectrometry (ESI-MS) detection has been developed for the study of zinc binding MTs' polymerization in tris(hydroxymethyl)-aminomethane (TRIS) acetate buffer at physiological pH. This hyphenated technique allows separating the different polymeric states of MTs by SEC, followed by on-line identification of the individual MT subisoforms in each polymeric peak by ESI-MS detection. Purified MT subisoforms (MT-2d and MT-2a), MT-2d and MT-2a mixture and rabbit liver MT complexes were investigated in the experiments to confirm the results obtained. From the results, both oxidative polymerization and non-oxidative oligomerization were found. The cystein-dependent oxidation results in the tetrameric peak as shown in the chromatograms of oxidized MT-2d, and stable dimeric and monomeric of MT were detected in this peak by MS. For the dimeric and trimeric peaks, different MT subisoforms were detected. In the five major subisoforms detected in rabbit liver MT complexes, MT-2a and MT-2c exist primarily as trimer, while MT-2e, MT-2d and MT-1a exist mainly as dimer. Our results suggest that in the three kinds of polymers, dimer, trimer and tetramer that were found in samples, the tetramer comes from the oxidation of MT molecular; for the dimer and trimer resulting from cystein independent oligomerization, they are closely associated with the charge of subisoform.

A method has been established for the separation and identification of metallothionein (MT) isoforms and sub-isoforms by liquid chromatography. A mixture of rabbit liver MT was separated on a DEAE A-25 weak anion-exchange chromatographic (AEC) column. Inductively coupled plasma-mass spectrometer (ICP-MS) was used for off-line detection of Zn in each fraction. The two main MT isoforms, MT-1 and MT-2, were isolated. The collected MT isoform fractions were then desalted by a homemade Sephadex G-25 size exclusion chromatographic (SEC) column. Afterwards, the sample was separated on a C18 reversed-phase column with UV detection at 210 nm. Different separation conditions were discussed and several sub-isoforms of MT were well separated at pH 2.0. The sub-isoforms were finally characterized by on-line HPLC-ESI-MS. The isoforms and sub-isoforms were well separated on a reversed-phase column under optimized chromatographic conditions and the detection results of ESI-MS were in agreement with the data found in literature. The method developed can be well used for the separation of metallothionein isoforms and sub-isoforms.

Hg concentrations, total metallothioneins (MTs) and metal-binding MTs in tissues of the rat after oral intake of HgCl2 were determined by the inductively coupled plasma-mass spectrometry (ICP-MS), a modifield mecurry saturarion assay and the size-exclusion chormatography (SEC) coupled with ICP-MS, respectively. The results indicated that Hg accumulation in all of the rat tissues, including kidney, liver, testicle, brain, heart, serum and blood plasma, in HgCl2 group significantly increased compared with that in the control group, especiallyin kidney,liver and testicle. A large amount of MTs was inducted in these exposed rat tissues, especially in their kidneys and livers. The metal species binding with MTs obtained with SECICP-MS showed that the fractions of Hg-MTs in both kidney and liver of exposed rat were significantly higher compared with those of the control. In addition, higher amounts of Cu-MTs observed in exposed rat liver and kidney demomstrated the effect of MTs in the homeostatis of Cu and in the detoxification of Hg.

Mammalian metallothioneins (MTs) usually have two major isoforms, MT-1 and MT-2, existing in tissues because of the genetic polymorphism. Metallothionein isoforms and sub-isoforms were characterized with a hyphenated technique of reversed-phase liquid chromatography (RPLC) coupled with electrospray ionization mass spectrometry (ESI-MS). The MT-isoform separation was carried out using a C8 narrow-bore column Vydac C8 (250 mm x 2.1 mm i.d., 5 microm, 30 nm). Buffer A was 5 mmol/L ammonium acetate in water (pH 6.0) and buffer B was 5 mmol/L ammonium acetate in methanol-water (1:1, v/v) (pH 6.0). A linear gradient elution was used by increasing buffer B from 10% to 37.5% within 40 min. The flow rate was set at 0.20 mL/min. The tentative formulas of MT isoforms and subisoforms were interpreted with the measured masses at the vicinity of apexes in total ion current chromatogram of ESI-MS and the reference data reported previously. The method was validated with the standard reference materials of MT-1 and MT-2. The results indicated that many MT species interpreted were in good agreement with the reference data reported. The MT sample from rabbit liver was measured with the established method. The results, of which the apo-MT was used by acidification of the sample for simplifying the mass spectra, showed that there were species of MT isoforms and subisoforms in rabbit livers.

COD determination based on ozone oxidation of alpha-naphthol combined with UV radiation (UV-O3) has been studied in the present work. Utilizing the phenomenon that luminol can be oxidized by the dissolved ozone to produce luminescence, we have established a new method of utilizing aqueous chemiluminescence to determine COD. The kinetics and mechanism of the ozonation reaction of alpha-naphthol have been investigated in order to gain a better understanding of the general applicability and limitation of the technique. Real world samples were analyzed and the results show that the relative error of COD(FI-CL) measurement for water samples was10%. Compared with the results of the conventional potassium permanganate method, the COD values of the FI-CL method are consistently higher (0-20% relative). The higher COD values suggest that the ozone-UV system is a more effective oxidation technique.