Arsenic speciation analysis in human saliva.
Studying arsenic species in saliva may increase our understanding of arsenic exposure, metabolism, and toxicity. Many studies on metabolism and toxicity of arsenic have relied on the analyses of biological fluids, including urine, blood, bile, and breast milk (10-20). No reports describe the detection of arsenic species in human saliva, however, although many chemical contaminants are metabolized and excreted to saliva (21).
A major challenge for detection of chemical contaminants in saliva is that the concentrations of the chemical contaminants are usually very low, often 1 to 2 orders of magnitude lower than in blood (4, 5).To achieve the sensitivity necessary for the detection of arsenic in saliva and to identify the arsenic species at ultratrace concentrations, we developed 2 highly specific techniques. We used inductively coupled plasma mass spectrometry (ICP-MS)  to detect arsenic species after separation by liquid chromatography (LC) and then applied electrospray tandem mass spectrometry (ESI-MS/MS) (16, 22) with multiple-reaction monitoring (MRM) to confirm the identification of arsenic species.
Materials and Methods
INSTRUMENTS AND REAGENTS
The instruments used in the study included an MDS SCIEX Applied Biosystems 4000Q-Trap MS/MS analyzer equipped with an electrospray ion-source, an Agilent 7500cs octopole reaction system ICP-MS, operated with the helium mode, and an Agilent 1100 series liquid chromatograph, equipped with an autosampler and column temperature control. Chromatographic separation was achieved with a Phenomenex reversed-phase ODS-3 column (150 x 4.6 mm, 3-[micro]m particle size) and a Hamilton PRP-X100 strong anion exchange column (150 x 4.1 mm) or a Hamilton PRP-X200 strong cation exchange column (150 x 4.1 mm).
We purchased arsenite ([As.sup.III]), arsenate ([As.sup.V]), and dimethylarsinic acid ([DMA.sup.V]) from Aldrich and of monomethylarsonic acid ([MMA.sup.V]) from Chem Service and prepared stock solutions (1000 mg/L) from these materials. We prepared working solutions by appropriate dilutions of the above stock solutions with deionized water. [MMA.sup.III] standard was prepared from the solid oxide (C[H.sub.3]AsO) and [DMA.sup.III] from the iodide [[(C[H.sub.3]).sub.2]AsI]. The precursors were prepared according to reported procedures (23) and were stored at 4[degrees]C or -20[degrees]C. Diluted solutions of the precursors were prepared fresh in deionized water. For quality control, we purchased the standard reference materials SRM1640 (Trace Elements in Natural Water) and SRM2670 (Trace Elements in Urine) from NIST. We used tetrabutylammonium hydroxide (Aldrich), malonic acid (Fisher), methanol (Fisher), and ammonium bicarbonate (Sigma) for preparation of the mobile-phase solutions, which were filtered through a 0.45-[micro]m membrane before use for LC separation.
This study was conducted according to the recommendations of the World Medical Association Declaration of Helsinki (24) for international health research. All participants gave written informed consent.
A saliva sample was collected from each of 32 volunteers (15 men and 17 women, age range 25 to 45 years) living in Edmonton, Canada. The concentration of arsenic in drinking water was <5 [micro]g/L. Saliva was also collected from 301 people living in Ba Men, Inner Mongolia, China, where the concentrations of arsenic in drinking water (ground water) are as high as 826 [micro]g/L. Demographic information on the participants has been reported for another biomarker study in the same population (25). The samples were collected at least 1 h after any food consumption; and before collection, participants rinsed their mouths at least 3 times to remove any food residue. The study participants provided saliva samples by spitting into 50-mL polyethylene centrifuge tubes. The samples were transported on dry ice via air courier and were stored at -20[degrees]C until analysis.
PRETREATMENT OF SALIVA FOR ARSENIC SPECIATION
The frozen saliva samples were thawed at room temperature and thoroughly vortex-mixed. We diluted the saliva 3-fold by pipetting 500 [micro]L saliva and 1mL deionized water into a 15-mL centrifuge tube. After vortex-mixing and ultrasonication, the dilute sample was filtered through a 0.45-[micro]m filter immediately before LC analysis.
DIGESTION OF SALIVA FOR TOTAL ARSENIC
We pipetted 300 [micro]L saliva and 1.0 mL concentrated nitric acid into a 5-mL glass vial. The mixture was then heated at 60[degrees]C for 12 h. The acid in the sample was evaporated by heating at 80[degrees]C until approximately 100 [micro]L of the solution remained. The sample was finally diluted to 1.5 mL with deionized water and analyzed for total arsenic by ICP-MS.
QUANTIFICATION OF ARSENIC SPECIES BY LC-ICP-MS
We carried out ion pair separation by use of the Phenomenex ODS-3 reversed-phase column with a mobile phase containing 5 mmol/L tetrabutylammonium, 5% methanol, and 3 mmol/L malonic acid (pH 5.6), with a flow rate of 1.2 mL/min. The column temperature was controlled at 48[degrees]C. [As.sup.III], [As.sup.V],[DMA.sup.V],[MMA.sup.V],[DMA.sup.III], and [MMA.sup.III] were separated in approximately 5 min. The eluent from the LC column was directly introduced to the inlet of the ICP nebulizer using polytetrafluoroethylene tubing and appropriate fittings (26). The ICP was operated at a radio frequency power of 1550 W. The flow rate of argon carrier gas was 0.8 L/min. Helium (3.6 mL/min) was used as the collision gas for the octopole reaction mode to reduce isobaric and polyatomic interference.
IDENTIFICATION OF ARSENIC SPECIES BY BOTH LC-ESI-MS/MS AND LC-ICP-MS
Initially, we operated ESI-MS/MS in the enhanced MS scan mode, in 1:1 methanol/water solution, to obtain MS/MS spectra of the arsenic species. The optimization of operating conditions was carried out using 200 [micro]g/L standard solutions of each arsenic species. The pH values of the [MMA.sup.V] and [DMA.sup.V] solutions were adjusted to 2.0-3.0 using formic acid, and [MMA.sup.V] and [DMA.sup.V] were monitored under the positive ionization mode. The optimal electrospray conditions were: curtain gas ([N.sub.2]) 1.1 L/min, ionspray voltage 5000 V, temperature 300[degrees]C, declustering potential 30 V, collision energy 30 V, and cell exit potential 26 V. [As.sup.III] and [As.sup.V] were monitored using the negative ionization mode, and the pH values of the [As.sup.III] and [As.sup.V] standard solutions was adjusted to 8.0-10. The optimal ionspray parameter settings for [As.sup.III] and [As.sup.V] were curtain gas ([N.sub.2]) 1.8 L/min, ionspray voltage -4500 V, temperature 300[degrees]C, and collision energy -20 V. The declustering potential was -100 V for [As.sup.III] and -130 V for [As.sup.V].
Because the mobile phase for the ion-pair chromatography was not compatible with ESI-MS/MS, a strong anion exchange column was used to separate arsenic species. The arsenic species were subsequently detected using both ICP-MS and ESI-MS/MS. Ammonium bicarbonate (20 mmol/L, pH 8.5) was used as the mobile phase at a flow rate of 0.8 mL/min. The LC effluent was split equally for ICP-MS and ESI-MS/MS detection. The flow to the ICP-MS was directly introduced to the nebulizer of ICP-MS. The flow to the ESI-MS/ MS was mixed with a solution containing methanol and either formic acid or ammonium hydroxide. Formic acid was introduced to the flow (adjusting the pH to 2.0-3.0) for the ESI-MS/MS detection of[MMA.sup.V] and [DMA.sup.V] under the positive ionization mode. Ammonium hydroxide was added to the flow (adjusting the pH to 8.0-10) for the ESI-MS/MS analysis of [As.sup.III] and [As.sup.V] under the negative ionization mode. ESI-MS/ MS was operated under the MRM mode. The characteristic ion transitions 139/91 and 139/121 were used for detecting [DMA.sup.V], 141/93 and 141/123 for [MMA.sup.V], 107/91 and 125/107 for [As.sup.III], and 141/123 for [As.sup.V].
DETERMINATION OF TOTAL ARSENIC BY FLOW INJECTION ICP-MS
We analyzed the acid-digested saliva samples for total arsenic using flow-injection ICP-MS. A 50-100 [micro]L sample was injected into a carrier stream of water at a flow rate of 1 mL/min, and was introduced to the ICP-MS.
SEPARATION AND IDENTIFICATION OF ARSENIC SPECIES USING LC WITH ICP-MS AND ESI-MS/MS DETECTION
Using ion-pair chromatography, we achieved the separation of 6 arsenic species, including [As.sup.III], [As.sup.V], [DMA.sup.V], [MMA.sup.V], [DMA.sup.III], and [MMA.sup.III], in approximately 5min. Speciation of saliva samples and samples enriched with arsenic species showed matching retention times for chromatographic peaks in saliva and the standards.
To confirm the identity of arsenic species in the samples, we further complemented the ICP-MS detection with ESI-MS/MS. Whereas ICP-MS provided detection of elemental arsenic (at m/z 75), ESI-MS/MS offered molecular information characteristic to each of the arsenic species. Therefore, we were able to achieve identification and quantification of arsenic species by using LC separation with both ICP-MS and ESI-MS/MS detection, as described below.
To obtain characteristic MS/MS information of the arsenic species, we initially carried out ESI-MS/MS analyses for each arsenic standard. MS/MS spectrum of [MMA.sup.V] showed the parent molecular ion [[[M.sup.+]H].sup.+] at m/z 141 and the characteristic product ions, including (C[H.sub.3])As(O)O[H.sup.+] at m/z 123, As[(OH).sub.2.sup.+] at m/z 109, HasO[H.sup.+] at m/z 93, (C[H.sub.3])As[H.sup.+] at m/z 91, As[H.sub.2.sup.+] at m/z 77, and [As.sup.+] at m/z 75. Similarly, the characteristic MS/MS spectrum of dimethylarsinic acid ([DMA.sup.V]) included the protonated molecular ion [[[M.sup.+]H].sup.+] at m/z 139 and its fragments at m/z 121, 109, 91, 89, 77, and 75. Mass spectra of [As.sup.III] and [As.sup.V] were obtained using negative ionization mode. The m/z 141 molecular ion and m/z 123, 93, and 77 fragment ions were characteristic of [As.sup.V]. The molecular and fragment ions for [As.sup.III] were m/z 125, 107, and 91, with m/z 107 being most abundant, corresponding to (O)As[O.sup.-].
Having characterized the MS/MS behavior of the arsenic species, we were able to select the characteristic MS/MS transitions for MRM detection of the particular arsenic species. Fig. 1 shows ion chromatography separation with ICP-MS monitoring of As at m/z 75 (Fig. 1a and 1b), MRM for [DMA.sup.V] transitions 139/91 (Fig. 1c) and 139/121 (Fig. 1d), and MRM for [MMA.sup.V] transitions 141/93 (Fig. 1e) and 141/123 (Fig. 1f). The matching of chromatographic retention times between the saliva sample (Fig. 1b) and arsenic standard solution (Fig. 1a) suggests the presence of [As.sup.III], [DMA.sup.V], [MMA.sup.V], and [As.sup.V] in the saliva sample. MRM monitoring of [DMA.sup.V] transitions at m/z 139/91 (Fig. 1c) and 139/121 (Fig. 1d) supports that the 2nd peak (3.8 min) is indeed[DMA.sup.V]. Likewise, the specific MRM of [MMA.sup.V] transition at m/z 141/93 (Fig. 1e) and 141/123 (Fig. 1f) supports the identity of[MMA.sup.V] (15.0 min) in the saliva sample.
Fig. 2 shows separation of arsenic in another saliva sample with ICP-MS detection of arsenic species (Fig. 2a), MRM monitoring for [As.sup.III] transitions at m/z 107/91 (Fig. 2b) and 125/107 (Fig. 2c), and MRM monitoring for a specific [As.sup.V] transition at m/z 141/123 (Fig. 2d). Retention times of the peaks detected by both ICP-MS (Fig. 2a) and ESI-MS/MS (Fig. 2b-d) match those of standards [As.sup.III] and [As.sup.V] (Fig. 1a). Cation exchange separation was also performed on selected samples to confirm the presence of [As.sup.III] and [As.sup.V] in the saliva sample.
DETERMINATION OF SALIVA ARSENIC CONCENTRATIONS
Having identified [As.sup.III], [As.sup.V], [MMA.sup.V], and [DMA.sup.V] species in saliva, we further quantified the concentrations of these arsenic species in saliva samples from volunteers (Table 1) and from a group of people who were exposed to varying concentrations of arsenic in drinking water (Table 2). Speciation analyses of saliva samples from 32 volunteers showed that [As.sup.III], [As.sup.V], and [DMA.sup.V] were the major arsenic species. [MMA.sup.V] was detected in 2 samples. Other arsenic species, if present, were below detection limits. Additional analyses of total arsenic, directly by ICP-MS without LC separation, showed consistent results with the sum of [As.sup.III], [As.sup.V], [MMA.sup.V], and [DMA.sup.V] concentrations obtained from the speciation analyses. The total arsenic concentration in the saliva samples ranged from 0.2 to 3.3 [micro]g/L.
[FIGURE 1 OMITTED]
[FIGURE 2 OMITTED]
The saliva samples from Inner Mongolia showed higher concentrations of arsenic (Table 2). The total arsenic concentrations ranged from 0.4 to 140 [micro]g/L. The major arsenic species were inorganic [As.sup.III] and [As.sup.V], accounting for 85% of total arsenic. Most of the samples (98%-99%) contained detectable [As.sup.III] and [As.sup.V], 80% of samples had detectable [MMA.sup.V], and 68% of samples had detectable [DMA.sup.V]. The mean percentages of individual arsenic species concentration over the mean total arsenic concentration were: [As.sup.III] 28%, [As.sup.V] 57%, [MMA.sup.V] 7%, and [DMA.sup.V] 4%. In addition, an unknown arsenic species was present in 20% of the samples and accounted for approximately 13% of total arsenic concentration in the saliva samples. The retention time (5.3 min) of this species was longer than that of [As.sup.V] and did not match that of any arsenic species available to us, including the 4 arsenic species shown in Fig. 1, plus [MMA.sup.III], [DMA.sup.III], arsenobetaine, trimethylarsine oxide (TMAO), and dimethylthiolarsenate (DMTA) produced by reacting [DMA.sup.V] with hydrogen sulfide ([H.sub.2]S) (27-29). We confirmed no retention time match between the unknown and the above arsenic species, using 3 chromatography systems: ion-pair chromatography (13), strong anion exchange (Figs. 1 and 2), and strong cation exchange (PRP-X200 column with 5 mmol/L pyridine and 5% methanol as mobile phase, pH 2.9). In addition, we used LC-ESI-MS/MS to analyze the saliva sample containing the unknown species. The ion transitions did not match those of our arsenic standards. The exact composition of the unknown arsenic species remains to be characterized.
Five samples had a detectable peak retention time identical to that of [MMA.sup.III], and 3 samples had a peak retention time matching that of[DMA.sup.III]. Because of the low concentration and the insufficient sample volume, however, we were not able to use MRM to confirm their identity in the saliva samples.
STANDARD REFERENCE MATERIALS
In the absence of standard reference material (SRM) for saliva arsenic, we analyzed Natural Water 1640 and 2 urine SRMs (SRM 2670 and CRM no. 18) (11, 13). Eleven repeat speciation analyses of Natural Water 1640 using LC-ICP-MS showed that the mean (SD) concentrations of [As.sup.III] and [As.sup.V] were 0.76 (0.05) [micro]g/L and 25.62 (0.61) [micro]g/L, respectively. Another 11 replicate analyses of this SRM by ICP-MS determined the mean (SD) total arsenic concentration to be 26.56 (0.36) [micro]g/L. These results are in excellent agreement with the certified value of total arsenic [26.67 (0.41) [micro]g/L, 99.6% accuracy]. Results from the analyses of urine SRM also showed good agreements with the certified values.
LIMITS OF DETECTION
With the LC-ICP-MS method, the limits of detection, defined as the concentration of arsenic that produced a chromatographic peak having intensity equal to 3 times the SD of the baseline noise (30), were 0.03 g/L for [As.sup.III],[MMA.sup.III], and[DMA.sup.V]; 0.05 g/L for[MMA.sup.V]; 0.08 g/L for [As.sup.V]; and 0.1 g/L for [DMA.sup.III]. Limits of quantification ranged from 0.1 g/L for [As.sup.III] and [DMA.sup.V] to 0.2 g/L for [MMA.sup.V], and 0.3 g/L for [As.sup.V].
RECOVERY AND COMPARISON BETWEEN THE SUM OF ARSENIC SPECIES AND TOTAL ARSENIC CONCENTRATIONS
To evaluate the possible effect of the saliva sample matrix on the separation and quantification of arsenicals, we measured recovery using the saliva samples from the volunteers separately enriched with each arsenic species. The enriched concentrations were 0.5-50 [micro]g/L for [As.sup.III] and [As.sup.V] and 0.5-5 [micro]g/L for the methylated arsenic species. The recoveries of [As.sup.III], [As.sup.V], [DMA.sup.V], and [MMA.sup.V] were 98%-100%. The recoveries of [MMA.sup.III] and[DMA.sup.III] were 82%-97%. The saliva sample matrix did not affect the separation of arsenic species, as shown by the identical retention times between arsenic standards and saliva samples (Figs. 1 and 2).
Each saliva sample was separately analyzed for individual arsenic species and for total arsenic concentrations. Results in Tables 1 and 2 show that the sum of arsenic species concentrations (obtained from LC-ICP-MS and LC-ESI-MS/MS) was in good agreement with the total arsenic concentrations (obtained from ICP-MS analyses without LC separation).
CORRELATION BETWEEN SALIVA ARSENIC SPECIES AND SKIN LESIONS
To evaluate the relationship between the presence of skin lesions and saliva arsenic concentrations, saliva arsenic measures were log (base 10) transformed and categorized into quartiles, and the adjusted odds ratios were calculated using logistic regression models. Table 3 summarizes the adjusted odds ratios for skin lesions by quartiles of arsenic concentrations. For all 4 arsenic species, with the increase of their concentrations in saliva, the number of people with skin lesions increases and the odds ratio for skin lesions also increases. The highest odds ratio (8.545) for skin lesions is associated with the highest quartile of [As.sup.III] concentration in saliva. Odds ratios for skin lesions associated with the highest quartiles of [As.sup.V], [MMA.sup.V], and [DMA.sup.V] concentrations were 3.246, 3.396, and 5.595, respectively. These associations were statistically significant (P <0.001).
Before this study, no data had been reported on arsenic concentration or speciation in human saliva. This report is the first to describe the detection of arsenic species in saliva and show the evidence of methyl arsenical metabolites, [MMA.sup.V] and [DMA.sup.V], as well as inorganic [As.sup.III] and [As.sup.V] in saliva.
The detection and identification of trace arsenic species in saliva are possible because of the coupling of highly sensitive and selective analytical techniques (LC-ICP-MS and LC-ESI-MS/MS). LC-ICP-MS enables selective quantification of arsenic species, and LC-ESI-MS/MS provides confirmation of the species identities. The mass spectra of arsenic species obtained in our experiment are in agreement with those previously obtained using a single quadrupole mass spectrometer (22).
Statistical analysis clearly showed that the prevalence of skin lesions increased with increasing concentrations of saliva arsenic species (Table 3). For example, among those with the lowest quartile of [As.sup.III] concentrations, the prevalence of skin lesions was 9.8%, but this increased to 39.2% among those in the highest quartile (adjusted odds ratio = 8.545, P=0). Similarly, the odds ratios for skin lesions increase with the concentrations of [As.sup.V], [MMA.sup.V], and [DMA.sup.V] (Table 3). For all saliva measures the trend with the prevalence of skin lesions was significant (P = 0.0001). Skin lesions are an important clinical symptom of chronic arsenic exposure and poisoning. The good correlation between the saliva arsenic concentration and the prevalence of skin lesions suggests that saliva arsenic can be a useful biomarker. Furthermore, correlation coefficients between arsenic species in saliva and total arsenic in drinking water were [As.sup.III] 0.470, [As.sup.V] 0.589, [MMA.sup.V] 0.663, and [DMA.sup.V] 0.485 (P <0.001 for all).
Arsenic in drinking water is mostly in inorganic forms, [As.sup.V] and [As.sup.III] (31-35). Various organic arsenic species are present in food (34-35). The presence of [MMA.sup.V] and [DMA.sup.V] in saliva samples suggests direct absorption of these arsenic species or methylation of inorganic arsenic (34-37). During this biomethylation process, monomethylarsonous acid ([MMA.sup.III]) and dimethylarsinous acid ([DMA.sup.III]) can be formed as intermediates (13, 14, 36, 37). These trivalent methylarsenic species are more toxic than the pentavalent arsenic species (36-40). A few saliva samples showed detectable peaks matching those of [MMA.sup.III] and [DMA.sup.III] standards by LC-ICP-MS analysis; however, we were not able to provide confirmation by ESIMS/ MS analysis because of the limited sample volume and very low concentrations of these species.
In conclusion, inorganic arsenicals and their methylation metabolites, [MMA.sup.V] and [DMA.sup.V], were quantified in saliva samples collected from people who were exposed to various concentrations of arsenic. The ability to quantify the individual arsenic species and the noninvasive collection of saliva samples make arsenic speciation in saliva a potentially useful biomonitoring approach for assessing human exposure to arsenic.
Grant/funding Support: This study was supported by the Canadian Water Network, the Metals in the Human Environment Research Network, the National Cancer Institute of Canada, Alberta Health and Wellness, and the International Consortium of Persistent Toxic Substances (Chinese Academy of Sciences). Dr. C. Yuan acknowledges the research funding support from North China Electric Power University.
Financial Disclosure: None declared.
Acknowledgments: This research has been reviewed by US EPA and approved for publication. Approval does not signify that the contents reflect views of the Agency or endorse the trade names mentioned. We thank Dr. W. R. Cullen (University of British Columbia, Canada) for providing the [MMA.sup.III] and [DMA.sup.III] standards.
Received May 17, 2007; accepted October 11, 2007.
Previously published online at DOI: 10.1373/clinchem.2007.092189
(1.) Barr DB, Wang RY, Needham LL. Biologic monitoring of exposure to environmental chemicals throughout the life stages: requirements and issues for consideration for the national children's study. Environ Health Perspect 2005;113:1083-91.
(2.) Hold KM, de Boer D, Zuidema J, Maes RAA. Saliva as an analytical tool in toxicology. Int J Drug Test 1995;1:1-36.
(3.) Denovan LA, Lu C, Hines CJ, Fenske RA. Saliva biomonitoring of atrazine exposure among herbicide applicators. Int Arch Occup Environ Health 2000;73:457-62.
(4.) Lu C, Irish RM, Fenske RA. Biological monitoring of diazinon exposure using saliva in an animal model. J Toxicol Environ Health Part A 2003;66: 2315-25.
(5.) Timchalk C, Poet TS, Kousba AA, Campbell JA, Lin Y. Noninvasive biomonitoring approaches to determine dosimetry and risk following acute chemical exposure: Analysis of lead or organophosphate insecticide in saliva. J Toxicol Environ Health Part A 2004;67:635-50.
(6.) Barbosa F, Tanus-Santos JE, Gerlach RF, Parsons PJ. A critical review of biomarkers used for monitoring human exposure to lead: advantages, limitations, and future needs. Environ Health Perspect 2005;113:1669-74.
(7.) Gonzalez M, Banderas JA, Baez A, Belmont R. Salivary lead and cadmium in a young population residing in Mexico City. Toxicol Lett 1997;93:55-64.
(8.) Silva MJ, Reidy JA, Samandar E, Herbert AR, Needham LL, Calafat AM. Detection of phthalate metabolites in human saliva. Arch Toxicol 2005; 79:647-52.
(9.) Gorodischer R, Koren G. Salivary excretion of drugs in children: Theoretical and practical issues in therapeutic drug monitoring. Dev Pharmacol Ther 1992;19:161-77.
(10.) Mandal BK, Suzuki KT. Arsenic round the world: a review. Talanta 2002;58:201-35.
(11.) Ma M, Le XC. Effect of arsenosugar ingestion on urinary arsenic speciation. Clin Chem 1998;44: 539-50.
(12.) Wang Z, Zhou J, Lu X, Gong Z, Le XC. Arsenic speciation in urine from acute promyelocytic leukemia patients undergoing arsenic trioxide treatment. Chem Res Toxicol 2004;17:95-103.
(13.) Le XC, Lu X, Ma M, Cullen WR, Aposhian HV, Zheng B. Speciation of key arsenic metabolic intermediates in human urine. Anal Chem 2000; 72:5172-7.
(14.) Del Razo LM, Styblo M, Cullen WR, Thomas DJ. Determination of trivalent methylated arsenicals in biological matrices. Toxicol Appl Pharmacol 2001;174:282-93.
(15.) Yoshida K, Inoue Y, Kuroda K, Chen H, Wanibuchi H, Fukushima S, Endo G. Urinary excretion of arsenic metabolites after long-term oral administration of various arsenic compounds to rats. J Toxicol Environ Health Part A 1998;54:179-92.
(16.) Inoue Y, Date Y, Sakai T, Shimizu N, Yoshida K, Chen H, Kuroda K, Endo G. Identification and quantification by LC-MS and LC-ICP-MS of arsenic species in urine of rats chronically exposed to dimethylarsinic acid (DMAA). Appl Organomet Chem 1999;13:81-8.
(17.) Gregus Z, Gyurasics A, Csanaky I. Biliary and urinary excretion of inorganic arsenic: monomethylarsonous acid as major biliary metabolite in rats. Toxicol Sci 2000;56:18-25.
(18.) Concha G, Vogler G, Nermell B, Vahter M. Low-level arsenic excretion in breast milk of native Andean women exposed to high levels of arsenic in the drinking water. Int Arch Occup Environ Health 1998;71:42-6.
(19.) Zhang X, Cornelis R, De J, Mees L, Vanderbiesen V, Cubber AD, Vanholder R. Accumulation of arsenic species in serum of patients with chronic renal disease. Clin Chem 1996;42:1231-7.
(20.) Francesconi KA, Tanggaard R, McKenzie CJ, Goessler W. Arsenic metabolites in human urine after ingestion of an arsenosugar. Clin Chem 2002;48:92-101.
(21.) Polkowska Z, Kozlowska K, and Namiesnik J. Biological fluids as a source of information on the exposure of man to environmental chemical agents. Crit Rev Anal Chem 2004;34:105-19.
(22.) Florencio MH, Duarte MF, Bettencourt AMM, Gomes ML, Vilas Boas LF. Electrospray mass spectra of arsenic compounds. Rapid Commun Mass Spectrom 1997;11:469-73.
(23.) Cullen WR, McBride BC, Manji H, Pickett AW, Reglinski J. The metabolism of methylarsine oxide and sulfide. Appl Organometal Chem 1989;3:71-8.
(24.) Mo J, Xia Y, Wade TJ, Schmitt M, Le XC, Dang R, Mumford JL. Chronic arsenic exposure and oxidative stress: OGG1 expression and arsenic exposure, nail selenium, and skin hyperkeratosis in Inner Mongolia. Environ Health Perspect 2006; 114:835-41.
(25.) World Medical Association. World Medical Association Declaration of Helsinki, Hong Kong. 1989.
(26.) Le XC, Lu X, Li X-F. Arsenic speciation. Anal Chem 2004;76:27A-33A.
(27.) Hansen HR, Raab A, Jaspars M, Milne BF, Feldmann J. Sulfur-containing arsenical mistaken for dimethylarinous acid [DMA(III)] and identified as a natural metabolite in urine: Major implications for studies on arsenic metabolism and toxicity. Chem Res Toxicol 2004;17:1086-91.
(28.) Raml R, Goessler W, Traar P, Ochi T, Francesconi KA. Novel thioarsenic metabolites in human urine after ingestion of an arsenosugar, 2',3'-dihydroxypropyl 5-deoxy-5-dimethylarsinoyl-[beta]-d-riboside. Chem Res Toxicol 2005;18:1444-50.
(29.) Fricke MW, Zeller M, Sun H, Lai VWM, Cullen WR, Shoemaker JA, Witkowski MR, Creed JT. Chromatographic separation and identification of products from the reaction of dimethylarsinic acid with hydrogen sulfide. Chem Res Toxicol 2005; 18:1821-9.
(30.) ACS Committee on Environmental Improvement. Guidelines for data acquisition and data quality evaluation in environmental chemistry. Anal Chem 1980;52:2242-9.
(31.) Terlecka E. Arsenic speciation analysis in water samples: a review of the hyphenated techniques. Environ Monit Assess 2005;107:259-84.
(32.) Watt C, Le XC. Arsenic speciation in natural waters. In Cai Y. (Ed). Biogeochemistry of Trace Elements. American Chemical Society Symposium Series, Washington, DC. 2002;11-32.
(33.) Gong Z, Lu X, Watt C, Wen B, He B, Mumford J, Ning Z, Xia Y, Le XC. Speciation analysis of arsenic in groundwater from Inner Mongolia with an emphasis on acid-leachable particulate arsenic. Anal Chim Acta 2006;555:181-7.
(34.) Cullen WR, Reimer KJ. Arsenic speciation in the environment. Chem Rev 1989;89:713-64.
(35.) National Research Council. Arsenic in Drinking Water. National Academy Press, Washington, DC. 1999.
(36.) Styblo M, Drobna Z, Jaspers I, Lin S, Thomas DJ. The role of biomethylation in toxicity and carcinogenicity of arsenic: a research update. Environ Health Perspect 2002;110(Suppl.):767-77.
(37.) Aposhian HV, Aposhian MM. Arsenic toxicology: Five questions. Chem Res Toxicol 2006;19:1-15.
(38.) Styblo M, Serves SV, Cullen WR, Thomas DJ. Comparative inhibition of yeast glutathione reductase by arsenicals and arsenothiols. Chem Res Toxicol 1997;10:27-33.
(39.) Styblo M, Del Razo LM, Vega L, Germolec DR, LeCluyse EL, Hamilton GA, Reed W, Wang C. Cullen WR, Thomas DJ. Comparative toxicity of trivalent and pentavalent inorganic and methylated arsenicals in rat and human cells. Arch Toxicol 2000;74:289-99.
(40.) Petrick JS, Ayala-Fierro F, Cullen WR, Carter DE, Aposhian HV. Monomethylarsonous acid ([MMA.sup.III]) is more toxic than arsenite in Chang human hepatocytes. Toxicol Appl Pharmacol 2000;163:203-7.
Chungang Yuan, [1,2] Xiufen Lu,  Nicole Oro,  Zhongwen Wang,  Yajuan Xia,  Timothy J. Wade,  Judy Mumford,  and X. Chris Le  *
 Analytical and Environmental Toxicology, Department of Laboratory Medicine and Pathology, University of Alberta, Edmonton, Alberta, T6G 2G3, Canada;  School of Environmental Sciences and Engineering, North China Electric Power University, Baoding 071003, Hebei Province, P. R. China;  Inner Mongolia Center for Endemic Disease Control and Research, Huhhot 010020, Inner Mongolia, P. R. China;  National Health and Environmental Effects Research Laboratory, Human Studies Division, U.S. Environmental Protection Agency, Research Triangle Park, NC 27711, U.S.A.
* Address correspondence to this author at: Department of Laboratory Medicine and Pathology, Faculty of Medicine and Dentistry, University of Alberta, 10-102 Clinical Sciences Building, Edmonton, Alberta, Canada T6G 2G3. Fax +1-780492-7800; e-mail email@example.com.
 Nonstandard abbreviations: ICP-MS, inductively coupled plasma mass spectrometry; LC, liquid chromatography; ESI-MS/MS, electrospray ionization tandem mass spectrometry; MRM, multiple reaction monitoring; [As.sup.III], arsenite; [As.sup.V], arsenate; [MMA.sup.V], monomethylarsonic acid; [MMA.sup.III], monomethylarsonous acid; [DMA.sup.III], dimethylarsinous acid; SRM, standard reference material.
Table 1. Summary of arsenic species concentration in saliva samples collected from volunteers in Edmonton, Canada. Mean Detectable Number of Species in concentration, range, Detectable saliva samples [micro] g/L [micro] g/L samples [As.sup.III] 0.30 ND-0.70 22 [As.sup.V] 0.29 ND-0.53 24 [DMA.sup.V] 0.68 ND-2.60 17 [MMA.sup.V] 0.14 ND-0.16 2 Sum 0.79 0.19-2.90 Total 0.91 0.19-3.30 Percentage Percent over total of samples arsenic, % Species in containing the saliva samples arsenic species, % Mean Range [As.sup.III] 69 20 0-86 [As.sup.V] 75 40 0-103 [DMA.sup.V] 53 23 0-78 [MMA.sup.V] 6 1 0-16 Sum - - Total Sum over total arsenic: 84% (70%-103%). ND, not detectable (below detection limits) Table 2. Summary of arsenic species concentration in saliva samples collected from Inner Mongolia. Mean Detectable Detectable Species in concentration, range, sample saliva samples [micro] g/L [micro] g/L No. [As.sup.III] 2.8 0.1-37.7 298 [As.sup.V] 8.1 0.3-120.3 295 [DMA.sup.V] 0.4 0.1-3.9 206 [MMA.sup.V] 0.8 0.1-6.0 240 Unknown 1.4 0.1-18.9 59 Sum 11.9 0.3-135 - Total 12.5 0.4-140 301 Percentage Percent over total of samples arsenic, % Species in containing the saliva samples arsenic species, % Mean Range [As.sup.III] 99 28 0.3-99 [As.sup.V] 98 57 15-99 [DMA.sup.V] 68 4 0.1-59 [MMA.sup.V] 80 7 0.2-24 Unknown 20 13 0.4-30 Sum - - - Total 100 Sum over total arsenic: 95% (79%-119%). Table 3. Adjusted odds ratios for skin lesions by quartile of salvia arsenic species. Skin lesions absent Concentration range, Species Quartile [micro] g/L Number % [As.sup.III] 1 0.09-0.75 61 32.8 2 0.76-1.52 49 26.3 3 1.53-3.06 45 24.2 4 3.09-37.7 31 16.7 [As.sup.V] 1 0.25-1.44 58 31.5 2 1.46-3.47 47 25.5 3 3.51-8.85 43 23.4 4 8.92-120 36 19.6 [MMA.sup.V] 1 0.08-0.23 43 31.2 2 0.24-0.49 41 29.7 3 0.50-0.93 30 21.7 4 0.94-5.97 24 17.4 [DMA.sup.V] 1 0.08-0.16 41 36.3 2 0.17-0.30 34 30.1 3 0.31-0.46 21 18.6 4 0.47-3.81 17 15.0 Skin lesions present Total Total Species Number % number % [As.sup.III] 10 9.8 71 24.7 24 23.5 73 25.3 28 27.5 73 25.3 40 39.2 71 24.7 [As.sup.V] 15 14.9 73 25.6 24 23.8 71 24.9 27 26.7 70 24.6 35 34.7 71 24.9 [MMA.sup.V] 13 14.0 56 24.2 20 21.5 61 26.4 27 29.0 57 24.7 33 35.5 57 24.7 [DMA.sup.V] 13 15.3 54 27.3 18 21.2 52 26.3 23 27.1 44 22.2 31 36.5 48 24.2 Species Odds ratio 95% CI p-value [As.sup.III] Reference (1.00) - - 2.653 (a) 1.108-6.353 0.029 3.711 (b) 1.548-8.898 0.003 8.545 (c) 3.523-20.725 0 [As.sup.V] Reference (1.00) - - 1.977 0.890-4.393 0.094 2.363 (a) 1.086-5.145 0.030 3.246 (b) 1.492-7.061 0.003 [MMA.sup.V] Reference (1.00) - - 1.697 0.696-4.140 0.245 3.121 (a) 1.309-7.439 0.010 3.396 (b) 1.427-8.081 0.006 [DMA.sup.V] Reference (1.00) - - 1.747 0.721-4.232 0.216 3.415 (b) 1.380-8.449 0.008 5.595 (c) 2.232-14.024 0.0002 Pearson chi2(3) for [As.sup.III], 28.1936; Pr 0.000; for [As.sup.V], 13.4167; Pr 0.004; for [MMA.sup.V], 16.7493; Pr 0.001; and for [DMA.sup.V], 20.0573; Pr 0.000. (a) P<0.05; (b) P<0.01; (c) P<0.001.
|Printer friendly Cite/link Email Feedback|
|Title Annotation:||Drug Monitoring and Toxicology|
|Author:||Yuan, Chungang; Lu, Xiufen; Oro, Nicole; Wang, Zhongwen; Xia, Yajuan; Wade, Timothy J.; Mumford, Jud|
|Date:||Jan 1, 2008|
|Previous Article:||Erythrocyte fatty acid composition and the metabolic syndrome: a National Heart, Lung, and Blood Institute GOLDN study.|
|Next Article:||Excretion of methamphetamine and amphetamine in human sweat following controlled oral metamphetamine administration.|