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Article(id=1218551258953204534, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1218551204993487072, articleNumber=null, orderNo=null, doi=10.16438/j.0513-4870.2018-0522, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1527868800000, receivedDateStr=2018-06-02, revisedDate=1532534400000, revisedDateStr=2018-07-26, acceptedDate=null, acceptedDateStr=null, onlineDate=1768454859722, onlineDateStr=2026-01-15, pubDate=1541952000000, pubDateStr=2018-11-12, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1768454859722, onlineIssueDateStr=2026-01-15, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1768454859722, creator=13701087609, updateTime=1768454859722, updator=13701087609, issue=Issue{id=1218551204993487072, tenantId=1146029695717560320, journalId=1189982191388893191, year='2018', volume='53', issue='11', pageStart='1761', pageEnd='1942', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1768454846857, creator=13701087609, updateTime=1768457167276, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1218560937590702204, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1218551204993487072, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1218560937590702205, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1218551204993487072, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=1879, endPage=1886, ext={EN=ArticleExt(id=1218551259561378677, articleId=1218551258953204534, tenantId=1146029695717560320, journalId=1189982191388893191, language=EN, title=Speciation analysis of trace mercury and arsenic in 31 kinds of animal drugs and discussion about the limit standards, columnId=1190335348761793317, journalTitle=Acta Pharmaceutica Sinica, columnName=ORIGINAL ARTICLES, runingTitle=null, highlight=null, articleAbstract=

The toxicity of heavy metals and harmful elements is close related to their speciation. In the present study, the methods for mercury and arsenic speciation analysis based on high-performance liquid chromatography conjunction with inductively coupled plasma mass spectrometry (HPLC-ICP-MS) were established and applied to the determination of 31 kinds of animal drugs, 29 of which were included in the Chinese Pharmacopeia (2015 edition). The results showed that the LODs for all the speciation were within 0.1-0.65 μg·kg-1, and the recoveries were within 86.9%-116.6% with the RSD of 1.49%-4.23%. Inorganic mercury (Hg2+) was detected in all the 87 batches of samples that came from 31 kinds of animal drugs, and the contents were 2.39-6567 μg·kg-1. Methylmercury (MeHg) was detected in 33 batches of samples that came from 12 kinds of animal drugs, and the contents were 2.83-319.7 μg·kg-1. Ethylmercury (EtHg) were detected in none of the samples. The detection rates of As(Ⅲ), As(Ⅴ), monomethylarsononous acid (MMA), dimethylarsinic acid (DMA), arsenobetaine (AsB) and arsenocholine (AsC) in the 31 batches of animal drugs was 96.77%, 100%, 45.16%, 90.32%, 93.55% and 22.58%, respectively. According to the toxic level of different speciation, the animal drugs with high risks of mercury were Agkistrodon, Bungarus Parvus, Zaocys, and Scolopendra; the animal drugs with high risks were Pheretima, Agkistrodon, Zaocys, and Aspongopus. This study can provide important evidence for the risk assessment, setting and revision of the limit standards of heavy metals and harmful elements.

, correspAuthors=Mei-hua YANG, authorNote=null, correspAuthorsNote=null, copyrightStatement=Copyright ©2018 Acta Pharmaceutica Sinica. All rights reserved., copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, authorCompany=null, fund=null, authors=null, authorsList=Jiao-yang LUO, Hao LIU, Shan-yong GU, Jian-jie WU, Mei-hua YANG), CN=ArticleExt(id=1218551260295381963, articleId=1218551258953204534, tenantId=1146029695717560320, journalId=1189982191388893191, language=CN, title=31种动物药中汞、砷元素形态残留分析与限量标准探讨, columnId=1190335348896011050, journalTitle=药学学报, columnName=研究论文, runingTitle=null, highlight=null, articleAbstract=

重金属与有害元素的毒性与其形态直接相关。本文建立了基于HPLC-ICP-MS的动物药中汞、砷形态分析方法，在此基础上检测了31种动物药（含2015版《中国药典》项下29个品种）中不同形态汞、砷的残留量。结果发现，不同形态汞、砷的LOQ为0.1~0.65 μg·kg-1，回收率为86.9%~116.6%，RSD为1.49%~4.23%。31种动物药共87个批次均检测到Hg2+，含量为2.39~6567 μg·kg-1；12种动物药共33个批次检测到MeHg，含量为2.83~319.7 μg·kg-1；所有的动物药样品中均未检测到EtHg。31种动物药批次中不同砷形态的检出率分别为：As（Ⅲ）：96.77%，As（Ⅴ）：100%，一甲基砷（MMA）：45.16%，二甲基砷（DMA）：90.32%，砷甜菜碱（AsB）：93.55%，砷胆碱（AsC）：22.58%。根据不同形态毒性强弱，Hg残留高风险的品种为：蕲蛇、金钱白花蛇、乌梢蛇、蜈蚣；As残留高风险的品种为：地龙、蕲蛇、乌梢蛇、九香虫。本研究可为动物药中重金属风险评估与限量标准制修订提供重要参考。

, correspAuthors=杨美华, authorNote=null, correspAuthorsNote=
* 杨美华, Tel:86-10-57833277, Fax:86-10-62896288, E-mail:
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Determination of Heavy Metals and Speciation Analysis of Mercury and Arsenic in Marine and Chinese Medicinal Herbs by ICP-MS (ICPMS用于海洋和中药材中重金属元素及其汞砷形态分析的研究)[D]. Jinan: Shan-dong University, 2008., articleTitle=null, refAbstract=null), Reference(id=1218970842223002150, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, doi=null, pmid=null, pmcid=null, year=null, volume=null, issue=null, pageStart=null, pageEnd=null, url=null, language=null, rfNumber=[19], rfOrder=18, authorNames=null, journalName=null, refType=null, unstructuredReference=Cao X. Identification of Arsenic and Arsenic Metabolites Based on Chemical Speciation Analysis (以形态分析为基础的砷及砷生物代谢过程与产物的表征研究)[D]. 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Acta Pharm Sin (药学学报), 2018, 53:432-438., articleTitle=null, refAbstract=null), Reference(id=1218970842613072440, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, doi=null, pmid=null, pmcid=null, year=null, volume=null, issue=null, pageStart=null, pageEnd=null, url=null, language=null, rfNumber=[23], rfOrder=22, authorNames=null, journalName=null, refType=null, unstructuredReference=Zuo TT, Li YL, Jin HY, et al. HPLC-ICP-MS speciation analysis and risk assessment of rsenic in Cordyceps sinensis[J]. Chin Med, 2018, 13:19., articleTitle=null, refAbstract=null)], funds=[Fund(id=1218970839941300654, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, awardId=2016-I2M-3-010, language=CN, fundingSource=中国医学科学院医学与健康科技创新工程(2016-I2M-3-010), fundOrder=null, country=null), Fund(id=1218970840046158261, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, awardId=2017-I2M-1-013, language=CN, fundingSource=中国医学科学院医学与健康科技创新工程(2017-I2M-1-013), fundOrder=null, country=null), Fund(id=1218970840125850042, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, awardId=201507004-4, language=CN, fundingSource=中医药行业科研专项(201507004-4), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1218970833993778136, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, xref=null, ext=[AuthorCompanyExt(id=1218970834010555355, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, companyId=1218970833993778136, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100193, China), AuthorCompanyExt(id=1218970834027332575, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, companyId=1218970833993778136, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=中国医学科学院、北京协和医学院药用植物研究所, 北京 100193)])], figs=[ArticleFig(id=1218970837932228896, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=EN, label=null, caption=null, figureFileSmall=8sbbOtW2QkK8EAY8b6tpZw==, figureFileBig=PWYB1CiT23PXvbymfoVAag==, tableContent=null), ArticleFig(id=1218970838024503592, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=CN, label=Figure 1, caption=

Chromatograms of (A) mix standards of mercury speciation, (B) a real sample of Scorpio, (C) mix standards of arsenic speciation, and (D) a real sample of Zaocys

, figureFileSmall=8sbbOtW2QkK8EAY8b6tpZw==, figureFileBig=PWYB1CiT23PXvbymfoVAag==, tableContent=null), ArticleFig(id=1218970838204858683, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
Time/min Mobile phase A/% Mobile phase B/%
0 100 0
1.5 100 0
4.0 0 100
7.5 0 100
8.0 100 0
10.0 100 0
), ArticleFig(id=1218970838313910593, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=CN, label=Table 1, caption=

Gradient elution program of arsenic speciation

, figureFileSmall=null, figureFileBig=null, tableContent=
Time/min Mobile phase A/% Mobile phase B/%
0 100 0
1.5 100 0
4.0 0 100
7.5 0 100
8.0 100 0
10.0 100 0
), ArticleFig(id=1218970838435545421, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
Speciation Regression equation Linear range
/μg·L−1		 R2 LOQ
/μg·L−1
Hg2+ y = 10 059 x − 320.09 0.2−10 0.999 8 0.4
MeHg y = 10 552 x − 756.3 0.2−10 0.999 9 0.25
EtHg y = 12 471 x − 298.18 0.2−10 0.999 9 0.65
AsB y = 1 544.9 x − 51.669 0.1−50 0.999 9 0.1
As(Ⅲ) y = 1 224 x − 65.845 0.2−100 0.999 9 0.2
DMA y = 1 481.9 x + 126.01 0.2−50 0.999 9 0.2
AsC y = 1 056.3 x − 151.01 0.5−50 0.999 9 0.4
MMA y = 1 212.1 x − 85.615 0.2−50 0.999 9 0.2
As(Ⅴ) y = 1 108.1 x + 1 767.6 0.2−100 0.999 6 0.2
), ArticleFig(id=1218970838582346073, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=CN, label=Table 2, caption=

Standard curves, linear ranges and limit of quantification. MeHg: Methyl mercury; EtHg: Ethyl mercury; As(Ⅲ): Trivalent arsenite; As(Ⅴ): Pentavalent arsenate; MMA: Monomethylarsononous acid; DMA: Dimethylarsinic acid; AsB: Arsenobetaine; AsC: Arsenocholine

, figureFileSmall=null, figureFileBig=null, tableContent=
Speciation Regression equation Linear range
/μg·L−1		 R2 LOQ
/μg·L−1
Hg2+ y = 10 059 x − 320.09 0.2−10 0.999 8 0.4
MeHg y = 10 552 x − 756.3 0.2−10 0.999 9 0.25
EtHg y = 12 471 x − 298.18 0.2−10 0.999 9 0.65
AsB y = 1 544.9 x − 51.669 0.1−50 0.999 9 0.1
As(Ⅲ) y = 1 224 x − 65.845 0.2−100 0.999 9 0.2
DMA y = 1 481.9 x + 126.01 0.2−50 0.999 9 0.2
AsC y = 1 056.3 x − 151.01 0.5−50 0.999 9 0.4
MMA y = 1 212.1 x − 85.615 0.2−50 0.999 9 0.2
As(Ⅴ) y = 1 108.1 x + 1 767.6 0.2−100 0.999 6 0.2
), ArticleFig(id=1218970838687203676, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
Analyte Initial content
/μg·kg−1		 Spiked
/μg·kg−1		 Found
/μg·kg−1		 Recovery
/%		 RSD
/%
Hg2+ 29.66 20 51.26 108.0 2.48
40 76.30 116.6 3.03
80 107.26 97.0 3.88
MeHg 47.31 20 65.47 90.8 3.66
40 85.19 94.7 4.01
80 127.63 100.4 3.73
AsB 110.85 50 166.1 110.5 2.85
100 209.55 98.7 3.48
200 295.65 92.4 4.15
As(Ⅲ) 175.14 50 227.39 104.5 2.47
100 268.94 93.8 3.18
200 366.54 95.7 1.49
DMA 20.48 10 30.24 97.6 1.78
20 40.96 102.4 3.45
40 59.16 96.7 2.94
AsC 6.87 5 11.6 94.6 2.76
10 15.56 86.9 4.16
20 26.21 96.7 3.74
MMA 6.08 5 10.805 94.5 2.65
10 14.83 87.5 3.59
20 25.72 98.2 1.86
As(Ⅴ) 23.79 10 34.14 103.5 3.45
20 45.53 108.7 2.89
40 63.11 98.3 4.23
), ArticleFig(id=1218970838821421414, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=CN, label=Table 3, caption=

Recoveries and precisions of the method in Scorpio and Zaocys samples. n = 3

, figureFileSmall=null, figureFileBig=null, tableContent=
Analyte Initial content
/μg·kg−1		 Spiked
/μg·kg−1		 Found
/μg·kg−1		 Recovery
/%		 RSD
/%
Hg2+ 29.66 20 51.26 108.0 2.48
40 76.30 116.6 3.03
80 107.26 97.0 3.88
MeHg 47.31 20 65.47 90.8 3.66
40 85.19 94.7 4.01
80 127.63 100.4 3.73
AsB 110.85 50 166.1 110.5 2.85
100 209.55 98.7 3.48
200 295.65 92.4 4.15
As(Ⅲ) 175.14 50 227.39 104.5 2.47
100 268.94 93.8 3.18
200 366.54 95.7 1.49
DMA 20.48 10 30.24 97.6 1.78
20 40.96 102.4 3.45
40 59.16 96.7 2.94
AsC 6.87 5 11.6 94.6 2.76
10 15.56 86.9 4.16
20 26.21 96.7 3.74
MMA 6.08 5 10.805 94.5 2.65
10 14.83 87.5 3.59
20 25.72 98.2 1.86
As(Ⅴ) 23.79 10 34.14 103.5 3.45
20 45.53 108.7 2.89
40 63.11 98.3 4.23
), ArticleFig(id=1218970838943056238, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
Certified reference material Analyte Found
/μg·kg−1		 Certified value
/μg·kg−1
Scallop Hg2+ 18.14 ± 0.54
GBW10024 MeHg 24.45 ± 0.95
EtHg ND
Total Hg 42.59 ± 1.49 40 ± 7
), ArticleFig(id=1218970839039525240, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=CN, label=Table 4, caption=

Contents of mercury speciation in certified reference scallop by HPLC-ICP-MS (n = 3)

, figureFileSmall=null, figureFileBig=null, tableContent=
Certified reference material Analyte Found
/μg·kg−1		 Certified value
/μg·kg−1
Scallop Hg2+ 18.14 ± 0.54
GBW10024 MeHg 24.45 ± 0.95
EtHg ND
Total Hg 42.59 ± 1.49 40 ± 7
), ArticleFig(id=1218970839131799934, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
Certified reference material Analyte Found
/μg·kg−1		 Certified value
/μg·kg−1
Lyophilized human urine As3+ 11.25 ± 0.10 11.4 ± 0.225
GBW09115 DMA 62.45 ± 2.3 64.5 ± 4.5
MMA 16.12 ± 0.50 15.45 ± 1.2
), ArticleFig(id=1218970839232463237, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=CN, label=Table 5, caption=

Contents of arsenic speciation in certified reference lyophilized human urine by HPLC-ICP-MS (n = 3, x ± s)

, figureFileSmall=null, figureFileBig=null, tableContent=
Certified reference material Analyte Found
/μg·kg−1		 Certified value
/μg·kg−1
Lyophilized human urine As3+ 11.25 ± 0.10 11.4 ± 0.225
GBW09115 DMA 62.45 ± 2.3 64.5 ± 4.5
MMA 16.12 ± 0.50 15.45 ± 1.2
), ArticleFig(id=1218970839316349325, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
Types Batch No Hg2+/μg·kg−1 MeHg/μg·kg−1
Batch 1 Batch 2 Batch 3 Batch 1 Batch 2 Batch 3
Bombyx Batryticatus 3 1 595 1 601 144.3 ND ND ND
Bubali Cornu 3 2.39 2.76 3.46 2.83 3.23 3.33
Margarita 3 5.07 7.56 59.25 ND ND ND
Faeces Trogopterori 3 529.1 1 562 42.58 ND ND ND
Scorpio 3 38.04 29.66 21.53 53.97 47.31 39.91
Pheretima 3 2 888 4 120 43.74 10.07 ND 16.56
Testudinis Carapax et Plastrum 3 10.19 7.94 22.08 ND ND ND
Eupolyphaga Steleophaga 3 728.2 738.1 21.18 ND ND ND
Cervi Cornu Pantotrichum 3 8.58 10.74 19.66 ND ND ND
Asini Corii Colla 3 4.44 6.34 3.65 ND ND ND
Cervi Cornu 3 1 604 778.2 9.90 ND ND ND
Ostreae Concha 3 12.91 33.21 10.50 ND ND ND
Cicadae Periostracum 3 3 036 3 406 127.6 ND ND ND
Margaritifera Concha 3 146.0 119.2 9.51 ND ND ND
Trionycis Carapax 3 15.93 32.09 10.59 ND ND ND
Gecko 3 7.63 24.13 11.96 21.59 91.97 30.22
Scolopendra 3 9.87 14.25 19.22 131.4 186.6 156.6
Hirudo 3 12.56 14.73 15.12 ND ND 17.12
Zaocys 3 5.25 8.23 4.50 97.49 104.5 47.80
Hippocampus 3 7.59 23.82 8.48 37.92 17.45 14.53
Gekko Japonicus Dumeril et Bibron 3 6 567 397.3 22.73 41.11 50.93 45.15
Agkistrodon 3 4.08 8.11 7.38 313.1 319.7 6.99
Syngnathus 3 8.32 8.52 7.05 7.75 9.37 11.82
Vespae Nidus 3 145.5 66.00 128.0 ND ND ND
Aspongopus 3 52.39 67.80 49.05 ND ND ND
Sepiae Endoconcha 3 56.36 20.56 10.15 ND ND ND
Bufonis Venenum 3 33.22 54.55 14.37 ND ND ND
Bungarus Parvus 3 48.87 72.71 15.74 158.1 225.4 137.8
Bovis Calculus 1 229.9 − − ND ND ND
Moschus 1 54.63 − − ND ND ND
Cordyceps 1 240.6 − − ND ND ND
), ArticleFig(id=1218970839408624020, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=CN, label=Table 6, caption=

The contents of mercury speciation in 31 animal medicines. ND: Not detected

, figureFileSmall=null, figureFileBig=null, tableContent=
Types Batch No Hg2+/μg·kg−1 MeHg/μg·kg−1
Batch 1 Batch 2 Batch 3 Batch 1 Batch 2 Batch 3
Bombyx Batryticatus 3 1 595 1 601 144.3 ND ND ND
Bubali Cornu 3 2.39 2.76 3.46 2.83 3.23 3.33
Margarita 3 5.07 7.56 59.25 ND ND ND
Faeces Trogopterori 3 529.1 1 562 42.58 ND ND ND
Scorpio 3 38.04 29.66 21.53 53.97 47.31 39.91
Pheretima 3 2 888 4 120 43.74 10.07 ND 16.56
Testudinis Carapax et Plastrum 3 10.19 7.94 22.08 ND ND ND
Eupolyphaga Steleophaga 3 728.2 738.1 21.18 ND ND ND
Cervi Cornu Pantotrichum 3 8.58 10.74 19.66 ND ND ND
Asini Corii Colla 3 4.44 6.34 3.65 ND ND ND
Cervi Cornu 3 1 604 778.2 9.90 ND ND ND
Ostreae Concha 3 12.91 33.21 10.50 ND ND ND
Cicadae Periostracum 3 3 036 3 406 127.6 ND ND ND
Margaritifera Concha 3 146.0 119.2 9.51 ND ND ND
Trionycis Carapax 3 15.93 32.09 10.59 ND ND ND
Gecko 3 7.63 24.13 11.96 21.59 91.97 30.22
Scolopendra 3 9.87 14.25 19.22 131.4 186.6 156.6
Hirudo 3 12.56 14.73 15.12 ND ND 17.12
Zaocys 3 5.25 8.23 4.50 97.49 104.5 47.80
Hippocampus 3 7.59 23.82 8.48 37.92 17.45 14.53
Gekko Japonicus Dumeril et Bibron 3 6 567 397.3 22.73 41.11 50.93 45.15
Agkistrodon 3 4.08 8.11 7.38 313.1 319.7 6.99
Syngnathus 3 8.32 8.52 7.05 7.75 9.37 11.82
Vespae Nidus 3 145.5 66.00 128.0 ND ND ND
Aspongopus 3 52.39 67.80 49.05 ND ND ND
Sepiae Endoconcha 3 56.36 20.56 10.15 ND ND ND
Bufonis Venenum 3 33.22 54.55 14.37 ND ND ND
Bungarus Parvus 3 48.87 72.71 15.74 158.1 225.4 137.8
Bovis Calculus 1 229.9 − − ND ND ND
Moschus 1 54.63 − − ND ND ND
Cordyceps 1 240.6 − − ND ND ND
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Types Batch No As(Ⅲ)
/μg·kg−1		 As(Ⅴ)
/μg·kg−1		 MMA
/μg·kg−1		 DMA
/μg·kg−1		 AsB
/μg·kg−1		 AsC
/μg·kg−1
Bombyx Batryticatus 3 132.7 ± 100.6 17.28 ± 5.77 ND < LOQ 3.81 ± 0.58 ND
Bubali Cornu 3 16.90 ± 4.02 7.16 ± 0.79 9.11 ± 0.73 45.67 ± 5.50 3.76 ± 0.75 ND
Margarita 3 ND 3.50 ± 0.51 ND < LOQ 1.80 ± 0.14 ND
Faeces Trogopterori 3 49.29 ± 41.29 19.73 ± 6.32 ND 28.34 ± 3.04 16.64 ± 2.40 ND
Scorpio 3 19.91 ± 4.41 11.67 ± 4.27 ND 2.47 ± 0.48 17.12 ± 7.10 ND
Pheretima 3 1 883 ± 391.5 431.8 ± 100.1 ND 32.12 ± 2.34 311.6 ± 12.70 ND
Testudinis Carapax et Plastrum 3 24.16 ± 23.99 3.10 ± 2.07 ND < LOQ 5.29 ± 0.88 ND
Eupolyphaga Steleophaga 3 178.2 ± 228.0 30.56 ± 8.23 ND 48.87 ± 5.65 37.96 ± 1.45 ND
Cervi Cornu Pantotrichum 3 4.39 ± 0.39 3.55 ± 0.98 ND < LOQ 2.64 ± 0.48 ND
Asini Corii Colla 3 19.69 ± 0.86 7.46 ± 1.77 ND < LOQ 1.59 ± 0.36 ND
Cervi Cornu 3 53.52 ± 45.26 6.00 ± 1.58 ND < LOQ 2.58 ± 0.35 ND
Ostreae Concha 3 7.38 ± 5.79 4.42 ± 1.60 ND ND 1.55 ± 0.77 ND
Cicadae Periostracum 3 59.45 ± 41.17 21.42 ± 1.46 3.11 ± 1.08 2.79 ± 2.50 11.84 ± 15.22 ND
Margaritifera Concha 3 18.94 ± 13.16 6.86 ± 5.27 < LOQ 4.24 ± 2.73 10.71 ± 4.19 ND
Trionycis Carapax 3 27.39 ± 9.39 5.98 ± 0.98 6.91 ± 1.80 4.02 ± 0.90 5.05 ± 1.06 ND
Gecko 3 7.68 ± 1.94 12.14 ± 4.11 3.62 ± 0.56 10.63 ± 2.97 23.86 ± 19.68 5.10 ± 2.94
Scolopendra 3 14.29 ± 2.39 9.85 ± 0.81 6.24 ± 1.84 3.78 ± 0.44 99.20 ± 21.94 ND
Hirudo 3 189.2 ± 73.12 10.29 ± 4.23 7.88 ± 3.68 40.61 ± 15.59 161.1 ± 187.5 19.51 ± 24.22
Zaocys 3 331.5 ± 202.6 24.95 ± 15.78 6.33 ± 1.79 30.84 ± 18.09 356.0 ± 406.0 120.8 ± 198.7
Hippocampus 3 12.94 ± 4.69 15.12 ± 3.46 < LOQ 6.80 ± 1.91 61.90 ± 31.94 9.29 ± 1.97
Gekko Japonicus Dumeril et Bibron 3 97.34 ± 33.27 15.50 ± 4.42 3.51 ± 1.64 27.24 ± 2.29 46.39 ± 4.58 8.45 ± 3.47
Agkistrodon 3 411.9 ± 85.47 15.45 ± 2.52 1.12 ± 0.64 2.58 ± 1.46 19.50 ± 12.13 ND
Syngnathus 3 10.94 ± 9.82 7.78 ± 6.01 ND 95.26 ± 32.74 2 016 ± 676.5 36.09 ± 6.60
Vespae Nidus 3 143.1 ± 173.4 20.63 ± 14.55 5.88 ± 0.62 13.32 ± 1.62 25.23 ± 3.15 ND
Aspongopus 3 275.5 ± 449.4 27.74 ± 28.38 ND 2.13 ± 0.23 6.28 ± 1.33 ND
Sepiae Endoconcha 3 36.63 ± 57.81 8.62 ± 8.71 ND 401.5 ± 122.9 1 257 ± 129.5 10.38 ± 0.75
Bufonis Venenum 3 10.94 ± 2.13 8.35 ± 4.11 ND ND ND ND
Bungarus Parvus 3 2.93 ± 1.13 4.90 ± 3.04 ND 62.28 ± 43.54 15.22 ± 17.66 ND
Bovis Calculus 1 5.79 4.44 ND ND ND ND
Moschus 1 11.72 19.00 5.16 3.65 9.92 ND
Cordyceps 1 165.31 219.6 83.12 4.97 11.53 ND
), ArticleFig(id=1218970839622533537, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1218551258953204534, language=CN, label=Table 7, caption=

The contents of arsenic speciation in 31 animal medicines. ND: Not detected

, figureFileSmall=null, figureFileBig=null, tableContent=
Types Batch No As(Ⅲ)
/μg·kg−1		 As(Ⅴ)
/μg·kg−1		 MMA
/μg·kg−1		 DMA
/μg·kg−1		 AsB
/μg·kg−1		 AsC
/μg·kg−1
Bombyx Batryticatus 3 132.7 ± 100.6 17.28 ± 5.77 ND < LOQ 3.81 ± 0.58 ND
Bubali Cornu 3 16.90 ± 4.02 7.16 ± 0.79 9.11 ± 0.73 45.67 ± 5.50 3.76 ± 0.75 ND
Margarita 3 ND 3.50 ± 0.51 ND < LOQ 1.80 ± 0.14 ND
Faeces Trogopterori 3 49.29 ± 41.29 19.73 ± 6.32 ND 28.34 ± 3.04 16.64 ± 2.40 ND
Scorpio 3 19.91 ± 4.41 11.67 ± 4.27 ND 2.47 ± 0.48 17.12 ± 7.10 ND
Pheretima 3 1 883 ± 391.5 431.8 ± 100.1 ND 32.12 ± 2.34 311.6 ± 12.70 ND
Testudinis Carapax et Plastrum 3 24.16 ± 23.99 3.10 ± 2.07 ND < LOQ 5.29 ± 0.88 ND
Eupolyphaga Steleophaga 3 178.2 ± 228.0 30.56 ± 8.23 ND 48.87 ± 5.65 37.96 ± 1.45 ND
Cervi Cornu Pantotrichum 3 4.39 ± 0.39 3.55 ± 0.98 ND < LOQ 2.64 ± 0.48 ND
Asini Corii Colla 3 19.69 ± 0.86 7.46 ± 1.77 ND < LOQ 1.59 ± 0.36 ND
Cervi Cornu 3 53.52 ± 45.26 6.00 ± 1.58 ND < LOQ 2.58 ± 0.35 ND
Ostreae Concha 3 7.38 ± 5.79 4.42 ± 1.60 ND ND 1.55 ± 0.77 ND
Cicadae Periostracum 3 59.45 ± 41.17 21.42 ± 1.46 3.11 ± 1.08 2.79 ± 2.50 11.84 ± 15.22 ND
Margaritifera Concha 3 18.94 ± 13.16 6.86 ± 5.27 < LOQ 4.24 ± 2.73 10.71 ± 4.19 ND
Trionycis Carapax 3 27.39 ± 9.39 5.98 ± 0.98 6.91 ± 1.80 4.02 ± 0.90 5.05 ± 1.06 ND
Gecko 3 7.68 ± 1.94 12.14 ± 4.11 3.62 ± 0.56 10.63 ± 2.97 23.86 ± 19.68 5.10 ± 2.94
Scolopendra 3 14.29 ± 2.39 9.85 ± 0.81 6.24 ± 1.84 3.78 ± 0.44 99.20 ± 21.94 ND
Hirudo 3 189.2 ± 73.12 10.29 ± 4.23 7.88 ± 3.68 40.61 ± 15.59 161.1 ± 187.5 19.51 ± 24.22
Zaocys 3 331.5 ± 202.6 24.95 ± 15.78 6.33 ± 1.79 30.84 ± 18.09 356.0 ± 406.0 120.8 ± 198.7
Hippocampus 3 12.94 ± 4.69 15.12 ± 3.46 < LOQ 6.80 ± 1.91 61.90 ± 31.94 9.29 ± 1.97
Gekko Japonicus Dumeril et Bibron 3 97.34 ± 33.27 15.50 ± 4.42 3.51 ± 1.64 27.24 ± 2.29 46.39 ± 4.58 8.45 ± 3.47
Agkistrodon 3 411.9 ± 85.47 15.45 ± 2.52 1.12 ± 0.64 2.58 ± 1.46 19.50 ± 12.13 ND
Syngnathus 3 10.94 ± 9.82 7.78 ± 6.01 ND 95.26 ± 32.74 2 016 ± 676.5 36.09 ± 6.60
Vespae Nidus 3 143.1 ± 173.4 20.63 ± 14.55 5.88 ± 0.62 13.32 ± 1.62 25.23 ± 3.15 ND
Aspongopus 3 275.5 ± 449.4 27.74 ± 28.38 ND 2.13 ± 0.23 6.28 ± 1.33 ND
Sepiae Endoconcha 3 36.63 ± 57.81 8.62 ± 8.71 ND 401.5 ± 122.9 1 257 ± 129.5 10.38 ± 0.75
Bufonis Venenum 3 10.94 ± 2.13 8.35 ± 4.11 ND ND ND ND
Bungarus Parvus 3 2.93 ± 1.13 4.90 ± 3.04 ND 62.28 ± 43.54 15.22 ± 17.66 ND
Bovis Calculus 1 5.79 4.44 ND ND ND ND
Moschus 1 11.72 19.00 5.16 3.65 9.92 ND
Cordyceps 1 165.31 219.6 83.12 4.97 11.53 ND
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31种动物药中汞、砷元素形态残留分析与限量标准探讨
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骆骄阳 , 刘好 , 谷善勇 , 吴建杰 , 杨美华 *
药学学报 | 研究论文 2018,53(11): 1879-1886
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药学学报 | 研究论文 2018, 53(11): 1879-1886
31种动物药中汞、砷元素形态残留分析与限量标准探讨
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骆骄阳, 刘好, 谷善勇, 吴建杰, 杨美华*
作者信息
  • 中国医学科学院、北京协和医学院药用植物研究所, 北京 100193

通讯作者:

* 杨美华, Tel:86-10-57833277, Fax:86-10-62896288, E-mail:
Speciation analysis of trace mercury and arsenic in 31 kinds of animal drugs and discussion about the limit standards
Jiao-yang LUO, Hao LIU, Shan-yong GU, Jian-jie WU, Mei-hua YANG*
Affiliations
  • Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100193, China
出版时间: 2018-11-12 doi: 10.16438/j.0513-4870.2018-0522
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重金属与有害元素的毒性与其形态直接相关。本文建立了基于HPLC-ICP-MS的动物药中汞、砷形态分析方法，在此基础上检测了31种动物药（含2015版《中国药典》项下29个品种）中不同形态汞、砷的残留量。结果发现，不同形态汞、砷的LOQ为0.1~0.65 μg·kg-1，回收率为86.9%~116.6%，RSD为1.49%~4.23%。31种动物药共87个批次均检测到Hg2+，含量为2.39~6567 μg·kg-1；12种动物药共33个批次检测到MeHg，含量为2.83~319.7 μg·kg-1；所有的动物药样品中均未检测到EtHg。31种动物药批次中不同砷形态的检出率分别为：As（Ⅲ）：96.77%，As（Ⅴ）：100%，一甲基砷（MMA）：45.16%，二甲基砷（DMA）：90.32%，砷甜菜碱（AsB）：93.55%，砷胆碱（AsC）：22.58%。根据不同形态毒性强弱，Hg残留高风险的品种为：蕲蛇、金钱白花蛇、乌梢蛇、蜈蚣；As残留高风险的品种为：地龙、蕲蛇、乌梢蛇、九香虫。本研究可为动物药中重金属风险评估与限量标准制修订提供重要参考。

动物药  /  重金属与有害元素  /  形态  /  风险评估  /  限量标准

The toxicity of heavy metals and harmful elements is close related to their speciation. In the present study, the methods for mercury and arsenic speciation analysis based on high-performance liquid chromatography conjunction with inductively coupled plasma mass spectrometry (HPLC-ICP-MS) were established and applied to the determination of 31 kinds of animal drugs, 29 of which were included in the Chinese Pharmacopeia (2015 edition). The results showed that the LODs for all the speciation were within 0.1-0.65 μg·kg-1, and the recoveries were within 86.9%-116.6% with the RSD of 1.49%-4.23%. Inorganic mercury (Hg2+) was detected in all the 87 batches of samples that came from 31 kinds of animal drugs, and the contents were 2.39-6567 μg·kg-1. Methylmercury (MeHg) was detected in 33 batches of samples that came from 12 kinds of animal drugs, and the contents were 2.83-319.7 μg·kg-1. Ethylmercury (EtHg) were detected in none of the samples. The detection rates of As(Ⅲ), As(Ⅴ), monomethylarsononous acid (MMA), dimethylarsinic acid (DMA), arsenobetaine (AsB) and arsenocholine (AsC) in the 31 batches of animal drugs was 96.77%, 100%, 45.16%, 90.32%, 93.55% and 22.58%, respectively. According to the toxic level of different speciation, the animal drugs with high risks of mercury were Agkistrodon, Bungarus Parvus, Zaocys, and Scolopendra; the animal drugs with high risks were Pheretima, Agkistrodon, Zaocys, and Aspongopus. This study can provide important evidence for the risk assessment, setting and revision of the limit standards of heavy metals and harmful elements.

animal drug  /  heavy metal and harmful element  /  speciation  /  risk assessment  /  limit standard
骆骄阳, 刘好, 谷善勇, 吴建杰, 杨美华. 31种动物药中汞、砷元素形态残留分析与限量标准探讨. 药学学报, 2018 , 53 (11) : 1879 -1886 . DOI: 10.16438/j.0513-4870.2018-0522
Jiao-yang LUO, Hao LIU, Shan-yong GU, Jian-jie WU, Mei-hua YANG. Speciation analysis of trace mercury and arsenic in 31 kinds of animal drugs and discussion about the limit standards[J]. Acta Pharmaceutica Sinica, 2018 , 53 (11) : 1879 -1886 . DOI: 10.16438/j.0513-4870.2018-0522
正文
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重金属与有害元素毒性和生物有效性不仅与其浓度相关, 还决定于其化学形态[1−3]。其中, 有机汞因其具有亲脂性和生物放大效应, 毒性大于无机汞, 例如甲基汞(MeHg)的毒性要远高于离子汞(ion-Hg), 并且MeHg具有极强的生物亲和力, 更容易进入体内, 主要侵犯中枢神经系统, 典型症状为末梢感觉错乱、视野收缩、运动性共济失调等, 甲基汞还可透过胎盘屏障, 对胎儿脑细胞发育造成严重损害[4]。日本水俣病事件就是由MeHg污染食物链引起的, 人体血液中MeHg的含量超过0.2 mg·kg−1就会出现中毒症状[5]。甲基汞已被列入2B类致癌物清单中, GB 2762-2017规定水产动物及其制品中MeHg的限量值为0.5 mg·kg−1, 联合国粮食及农业组织和世界卫生组织(FAO/WHO)联合食品添加剂专家委员会(JECFA)规定MeHg的暂定每周可耐受摄入量(PTWI)为1.6 μg·kg−1体重, 美国环境保护署(EPA)规定人体每日摄入MeHg的参考剂量(RfD)为0.1 μg·kg−1。对于砷(As), 无机As的毒性大于有机砷, 亚砷酸盐化合物的可溶性、迁移性和毒性更高[6], 三价砷[As(Ⅲ)]的毒性是五价砷[As(Ⅴ)]的60倍, 是一甲基砷(MMA)的70倍[7]; 而有机态的As中, MMA的毒性要强于其他的有机态As, 砷甜菜碱(AsB)、砷胆碱(AsC)等则基本无毒, 以砷化合物的半数致死量LD50计, 其毒性由大到小依次为AsH3 > As(Ⅲ) > As(Ⅴ) > MMA > DMA > TMAO > AsC > AsB > As[8, 9]。重金属与有害元素主要通过生物链的富集作用进入动物或人体, 其富集倍数可高达1×106~1×107[10, 11]。动物药是来源于动物的药物, 为动物的全体﹑器官或组织等, 与植物药相比, 其在生物链中富集有机态元素的途径更广, 残留问题也变得越来越突出[12], 应给予足够的重视。
然而, 目前动物药的重金属与有害元素残留研究并不系统和深入, 品种涉及不全面, 且主要围绕残留总量开展研究[13], 2015版《中国药典》收录了阿胶、水蛭、蜂胶、牡蛎、蛤壳、珍珠、昆布、海藻、海螵蛸等动物药的重金属与有害元素限量标准[14]。虽有风险评估工作, 但基本是参考当前的国家或国际标准关于总残留量限量的初步分析[15]。据统计, 目前动物药蟾酥、蜈蚣检测过AsB和AsC[16−19], 全蝎检测过6种As元素形态; 尚未发现动物药中Hg元素形态检测的报道, 部分有海产品中Hg元素形态研究。另外, 有少量文献报道了海水[20]、海鱼[21]、白鲸和北极熊中Hg形态[19, 20]。综上所述, 动物药中重金属形态研究及质量控制处于初级阶段, 还有很长的路要走。本文拟对常用动物中As、Hg元素形态残留情况进行系统研究, 为科学的风险评估及质量控制提供基础数据。
材料与方法
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材料    31种动物药收集于药材市场、各大药店等销售点, 冷藏运输至实验室及时分批打粉并处理。
仪器与试剂    Thermo U-3000高效液相色谱仪(HPLC); 美国iCAP-Q电感耦合等离子体质谱仪(ICP-MS); 美国Millipore超纯水仪(Milli-Q); 德国Christ Alpha 2-4 LD plus实验室型冻干机; 凯达牌TGL16M台式高速冷冻离心机。Hg元素溶液标准物质[GBW(E) 080124, 100 μg·mL−1]、MeHg溶液标准物质(GBW08675, 65.0 ± 2.5 μg·mL−1)、EtHg溶液标准物质[GBW(E) 081524, 70.6 ± 2.5 μg·mL−1]、亚砷酸根[As(Ⅲ), GBW08666, 以As计, 75.7 μg·g−1]、砷酸根[As(Ⅴ), GBW08667, 以As计, 17.5 μg·g−1]、一甲基砷(MMA, GBW08668, 以As计, 25.1 μg·g−1)、二甲基砷(DMA, GBW08669, 以As计, 52.9 μg·g−1)、砷甜菜碱(AsB, GBW08670, 以As计, 38.8 μg·g−1)、砷胆碱(AsC, GBW08671, 以As计, 28.0 μg·g−1)、扇贝成分分析标准物质(GBW10024)、冻干人尿中砷形态成分标准物质(GBW09115), 均购自中国计量科学研究院。乙酸铵(≥99.99%, trace metals basis)、2-巯基乙醇(≥99.0%)、碳酸铵(99.999%, trace metals basis)购自美国Sigma公司; L-半胱氨酸(99%)购自上海阿拉丁生化科技股份有限公司; 盐酸(优级纯)购自国药集团化学试剂有限公司; 硝酸(优级纯)购自美国Fisher公司。
色谱条件    Hg形态: Agilent ZORBAX SB-C18 (250 mm × 4.6 mm, 3.5 μm)分析柱, Agilent ZORBAX SB-C18 (12.5 mm × 4.6 mm, 5 μm)保护住, 流动相:甲醇−0.01 mol·L−1乙酸铵溶液(含0.12% L-半胱氨酸) (8:92), 等度洗脱, 流速0.8 min·mL−1。As形态:离子色谱分析柱Dionex IonPacTM Analytical AS7 (4 mm × 250 mm), 保护柱Dionex IonPacTM Guard AS7 (4 mm × 50 mm), 流动相A 2 mmol·L−1碳酸铵溶液, 流动相B 100 mmol·L−1碳酸铵溶液, 流速1.0 mL·min−1, 梯度洗脱程序见表 1。
质谱条件    等离子体射频功率: 1 550 W, 冷却气Ar流速: 14 L·min−1, 雾化气Ar流速: 1.09 L·min−1, 辅助气Ar流速: 0.8 L·min−1, 汞形态测定为STD模式; 砷形态测定为KED模式, 碰撞气He流速: 5.075 mL·min−1, 蠕动泵转速: 40 r·min−1, 监测质量数202Hg和75As。
Hg混合标准溶液    精密称取MeHg和EtHg标准溶液适量(甲醇溶液), 以及Hg2+标准溶液(3%硝酸溶液)适量, 加8%甲醇−92%水(v/v)配制成3种Hg形态浓度均为1 μg·mL−1的混合母液, 以8%甲醇−92%水(v/v)分别稀释成质量浓度为10.00、8.00、5.00、4.00、2.00、0.50和0.20 μg·L−1的Hg形态系列溶液。
As混合标准溶液    分别精密称取MMA、DMA、As(Ⅲ)、As(Ⅴ)、AsB、AsC标准溶液(水溶液)适量, 加超纯水配制成As形态质量浓度均为1 μg·mL−1的混合母液, 以超纯水逐级稀释成质量浓度为50.00、20.00、10.00、5.00、1.00和0.50 μg·L−1等的As形态系列溶液。
供试品溶液的制备
Hg形态    取动物药粉末约0.5 g, 精密称定, 加入提取液(0.12% L-半胱氨酸, 0.10% 2-巯基乙醇, 0.10% HCl溶液) 5 mL, 涡旋混匀静置30 min, 35 ℃ 40 kHz超声提取30 min, 5 000 r·min−1离心5 min, 取上清液0.5 mL于冻干机中冻干, 加0.5 mL流动相溶液复溶, 加至0.5 mL-3 kDa Millipore超滤离心管14 000 r·min−1离心20 min, 滤液供HPLC-ICP-MS进样检测。对于超出线性上限样品, 选择超滤后的样品溶液, 加入流动相稀释至适当倍数后再次进样分析, 计算含量。
As形态    取动物药粉末约0.5 g, 精密称定, 加入纯水5 mL, 涡旋混匀静置30 min, 70 ℃ 40 KHz超声提取30 min, 5 000 r·min−1离心5 min, 收集上清液, 加至0.5 mL-3 kDa Millipore超滤离心管14 000 r·min−1离心20 min, 滤液供HPLC-ICP-MS进样检测。
标准样品的制备
扇贝    取扇贝标准物质约0.5 g, 精密称定, 参照供试品溶液制备项下Hg形态方法制备。
冻干人尿    取冻干人尿As形态标准物质(冻干品, 西林瓶装), 加5.00 mL超纯水溶解, 过0.22 μm滤膜, 待用。
结果
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1 方法学考察
1.1 标准工作曲线与检出限
取1 μg·mL−1 As、Hg形态混合标样母液, 分别加8%甲醇−92%水、超纯水, 逐级稀释各形态系列溶液。以As、Hg各形态的峰面积为纵坐标, 以浓度为横坐标, 绘制标准曲线。结果表明As、Hg各元素形态的标准曲线线性关系良好, R2值均大于0.999, 3种Hg形态的线性范围为0.2~10 μg·L−1, 5种As形态的线性范围为0.1~100 μg·L−1, 混标色谱图见图 1A、C, 线性考察结果见表 2。
在进样量为10 μL, 流速为0.8 min·mL−1的条件下, 采用逐级稀释法对标准溶液进行稀释, 参考2015版《中国药典》项下药品质量标准分析方法验证指导原则, 以信噪比为10:1时相对应浓度确定定量限, 测定Hg2+、MeHg、EtHg的定量限分别为0.4、0.25、0.65 μg·L−1; 在进样量为10 μL, 流速为1.0 min·mL−1的条件下, 采用逐步稀释法测定AsB、As(Ⅲ)、DMA、AsC、MMA、As(Ⅴ)的定量限分别为0.1、0.2、0.2、0.4、0.2和0.2 μg·L−1。
1.2 仪器精密度
取含浓度为1 μg·L−1 Hg和5 μg·L−1 As的混合标准溶液, 连续进样6次, 分别以As、Hg各形态峰面积值计算RSD值。各形态的RSD值均小于2%, 说明仪器精密度良好。
1.3 重复性
分别取全蝎、乌梢蛇粉末约0.5 g, 精密称定, 平行操作6份, 分别按供试品溶液中Hg形态和As形态的制备方法操作, 按照上述色谱条件和质谱条件进行测定, 计算Hg、As元素形态含量。结果表明Hg2+、MeHg的含量均值分别为46.12、45.12 μg·kg−1, RSD分别为5.31%、6.16%; AsB、As(Ⅲ)、DMA、AsC、MMA、As(Ⅴ)的含量均值分别为102.48、170.12、20.18、5.77、6.22和23.15 μg·kg−1, RSD分别为4.73%、3.99%、6.64%、5.03%、2.41%和3.84%。
1.4 加样回收率
取重复性实验所用全蝎、乌梢蛇样品粉末各约0.5 g (已计算Hg、As元素形态含量), 精密称定, 置于离心管中, 分别加入3个水平的Hg、As标准溶液(表 3), 按照供试品溶液的制备方法及上述色谱和质谱条件进行测定, 计算加样回收率。结果表明Hg2+、MeHg在20、40和80 μg·kg−1的添加水平时的加样回收率均值范围为90.8%~116.6%; AsB、As(Ⅲ)在50、100和200 μg·kg−1的添加水平时的加样回收率均值范围为92.0%~117.4%; DMA、As(Ⅴ)在10、20和40 μg·kg−1的添加水平时的加样回收率均值范围为96.7%~108.7%; AsC、MMA在5、10和20 μg·kg−1的添加水平时的加样回收率均值范围为86.9%~98.2% (表 3)。
2 标准样品校验
取扇贝标准物质约0.5 g, 精密称定, 参照供试品溶液制备项下Hg形态方法制备。结果表明Hg2+、MeHg的平均检出值分别为18.14和24.45 μg·kg−1, 3种常见形态的总量为(42.59 ± 1.49)μg·kg−1, 在标定值(40 ± 7) μg·kg−1范围内(表 4)。
取冻干人尿As形态标准物质, 按照说明加超纯水溶解, 过膜检测, 结果表明As(Ⅲ)、DMA、MMA的检出值分别为11.25 ± 0.10、62.45 ± 2.3、16.12 ± 0.50 μg·kg−1, 在标定值的范围内(表 5)。
以上结果表明所建立的方法适合于动物药中As、Hg元素形体的定量分析。
3 样品分析
采用上述建立的方法和操作步骤, 采用HPLC-ICP-MS法对31种动物药共87个批次的As、Hg元素形态进行了测定, 结果见表 6、表 7和图 1B, D。
牛黄、麝香和冬虫夏草收集到1批, 其他品种各收集到3批。结果发现, 31种动物药共87个批次均检测到Hg2+, 含量为2.39~6567 μg·kg−1, 其中地龙、蝉蜕、僵蚕、鹿角的检出值最高, 冬虫夏草和牛黄次之; 12种动物药共33个批次检测到MeHg, 含量为2.83~319.7 μg·kg−1, 其中蕲蛇、金钱白花蛇、蜈蚣、乌梢蛇的检出值较高。EtHg均未检出(表 6)。其中有部分样品的单批检出值与另外两批差异较大, 提示重金属污染不仅与品种相关, 也与其生长环境相关。
31种动物药批次中不同砷形态的检出率分别为: As(Ⅲ): 96.77%, As(Ⅴ): 100%, MMA: 45.16%, DMA: 90.32%, AsB: 93.55%, AsC: 22.58%。其中, As(Ⅲ)检出值最高的品种为地龙, 其次为蕲蛇、乌梢蛇、水蛭、僵蚕; As(Ⅴ)与MMA检出值最高的品种为地龙; DMA检出值最高的品种为海螵蛸、海龙, 其次为金钱白花蛇、土鳖虫、水蛭; AsB检出值较高的是海龙、海螵蛸; AsC检出值最高的品种为海龙、海螵蛸, 其次为地龙、乌梢蛇、蜈蚣。结果见表 7。
讨论
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本文对常用动物药的Hg、As元素形态进行了分析, 结果表明重金属总量与形态并非呈简单的正相关。以Hg元素为例, 大部分动物药品种以Hg2+为主(如地龙、蝉蜕), 少部分则以MeHg为主(如蛇类动物药)。蕲蛇的Hg2+含量低于10 μg·kg−1, 但其中MeHg含量高达300 μg·kg−1, 提示该品种的风险较高, 与地龙(Hg2+ > 4 000 μg·kg−1, MeHg < 20 μg·kg−1)相比更有控制必要。另外, 地龙中As(Ⅲ)和As(Ⅴ)含量高, 而海洋动物药中AsB与AsC含量高。由于不同形态的重金属毒性可能相差极大, 单以总量控制的模式存在明显的弊端。
中药的风险评估工作刚刚起步, 重金属的风险评估也逐渐有报道。Zhou等[22]采用人工胃肠消化模型, 测定重金属在全粉及煎煮入药方式下的转移率, 并利用靶标危害系数法(target hazard quotient, THQ)对黄连的重金属风险进行风险评估, 该评估是基于5种重金属的总摄入量及转移率, 结果黄连重金属含量处于安全范围, 不会对人体产生明显危害。Zuo等[23]对冬虫夏草中砷进行了风险评估, 该评估是基于WHO推荐的无机As的暂定每周耐受摄入量(provisional tlerable weekly intake, PTWI), 结合危害指数(hazard index, HI)、癌症风险(cancer risk, CR)、THQ等进行风险评估, 进而提出人群建议消费率2月/年时可将健康风险降至最低。
笔者认为中药材重金属风险评估首先应明确评估目的, 收集相关信息和数据, 可先进行保守评估, 如评估结论是风险极低, 则无需进一步评估; 相反, 对于风险高的品种, 应针对其毒性强的存在形态进行评估, 对中药材(如动物药)的每日服用量、服用天数等进行系统的调研和统计(如中国居民中药消费情况调查), 并选择合适的模型进行风险评估。此外, 在进行中药材中重金属风险评估时, 不应忽略日常膳食的重金属暴露, 明确留给中药的暴露空间。
最后, 关于重金属与有害元素现行的限量标准, 中国药典2015版共收载动物药46种, 部分动物药如水蛭、牡蛎、海藻、海螵蛸等收录了重金属总限量[14], 但并未收录Hg、As残留水平很高的地龙、蝉蜕、僵蚕等品种。更为重要的是, 现行的标准控制模式对于高毒形态的残留无法控制, 如蕲蛇、金钱白花蛇、蜈蚣等, 而对于这类动物药品种需要针对某种或某一类高毒形态进行评估(如MeHg、无机As), 计算最大限量理论值并结合中药的污染特点对理论值进行修订, 制定适合我国当前国情的动物药重金属形态限量标准。
基金
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  • 中国医学科学院医学与健康科技创新工程(2016-I2M-3-010)
  • 中国医学科学院医学与健康科技创新工程(2017-I2M-1-013)
  • 中医药行业科研专项(201507004-4)
文献
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2018年第53卷第11期
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doi: 10.16438/j.0513-4870.2018-0522
  • 接收时间：2018-06-02
  • 首发时间：2026-01-15
  • 出版时间：2018-11-12
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  • 收稿日期：2018-06-02
  • 修回日期：2018-07-26
基金
中国医学科学院医学与健康科技创新工程(2016-I2M-3-010)
中国医学科学院医学与健康科技创新工程(2017-I2M-1-013)
中医药行业科研专项(201507004-4)
作者信息
    中国医学科学院、北京协和医学院药用植物研究所, 北京 100193

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* 杨美华, Tel:86-10-57833277, Fax:86-10-62896288, E-mail:
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2种不同金属材料的力学参数
科
Family		 属数
Number of
genus		 种数
Number of
species		 占总种数比例
Percentage of
total species (%)		 属
Genus		 种数
Number of
species		 占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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