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Article(id=1240945601282633762, tenantId=1146029695717560320, journalId=1205117023404326918, issueId=1240945593548337937, articleNumber=null, orderNo=null, doi=10.16155/j.0254-1793.2024-0212, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1711728000000, receivedDateStr=2024-03-30, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1773794087000, onlineDateStr=2026-03-18, pubDate=1732982400000, pubDateStr=2024-12-01, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1773794087000, onlineIssueDateStr=2026-03-18, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1773794087000, creator=13701087609, updateTime=1773794087000, updator=13701087609, issue=Issue{id=1240945593548337937, tenantId=1146029695717560320, journalId=1205117023404326918, year='2024', volume='44', issue='11', pageStart='1827', pageEnd='2010', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1773794085156, creator=13701087609, updateTime=1773796488495, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1240955673937236736, tenantId=1146029695717560320, journalId=1205117023404326918, issueId=1240945593548337937, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1240955673937236737, tenantId=1146029695717560320, journalId=1205117023404326918, issueId=1240945593548337937, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=1983, endPage=1991, ext={EN=ArticleExt(id=1240945601576235068, articleId=1240945601282633762, tenantId=1146029695717560320, journalId=1205117023404326918, language=EN, title=Determination of 17 chemicals in medical cold compress by solid phase extraction-ultra performance liquid chromatography-tandem mass spectrometry*, columnId=1206272757852074373, journalTitle=Chinese Journal of Pharmaceutical Analysis, columnName=Safety Monitoring, runingTitle=null, highlight=null, articleAbstract=
Objective:

To establish a method for the determination of 17 kinds of anti-allergic compounds in medical cold compress by solid phase extraction-ultra performance liquid chromatography-tandem mass spectrometry（SPE-UPLC-MS/MS）. Common medical cold compress matrices were suitable for this new method.

Methods:

Three pretreatment methods were compared：prime pass-through column purification，SPE purification including HLB and MCX solid-phase extraction. The pretreatment was optimized by focusing on the kind of solid-phase extraction，the type and volume of elution solvents. The analytes were extracted using methanol and then mixed with water at the ratio of 1:9 in volume，purified with an Agilent Bond Elut HLB column. Waters ACQUITY UPLC BEH C18（100 mm×2.1 mm，1.8 μm）column was used with 0.1% formic acid(with 0.5 mmol·L-1 ammonium acetate)-acetonitrile as mobile phases by gradient elution for separating 17 compounds. Detection was carried out by positive and negative electrospray ionization（ESI）mass spectrometer in multiple reaction monitor (MRM) mode.

Results:

All the 17 kinds of compounds showed good linearity in their reasonable ranges（r）>0.99，the limit of detection （LOD） and quantification（LOQ）were 0.01-13.32 μg·g-1 and 0.02-26.64 μg·g-1，respectively. The average recoveries of at three spiked levels were in range of 77.7%-108.5%，with RSDs （n=6） were 0.62%-4.7%. The content range of diclofenac sodium in positive samples was 0.09-27.55 mg per tablet.

Conclusion:

The method is simple，sensitive and reliable，and is suitable for determination of 17 illegally add chemical drugs in medical cold compress. Moreover，the method might provide technological support for the detection of illegal additions in medical cold compress.

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目的：

采用固相萃取-超高效液相色谱-串联质谱（SPE-UPLC-MS/MS）法建立医用冷敷贴中17个化学物质的检测方法，该法适用于常见医用冷敷贴基质。

方法：

重点考察了固相萃取柱类型、洗脱溶剂种类及溶剂用量。样品用甲醇超声提取后，与水以1:9的比例混合，经Agilent Bond Elut HLB固相萃取柱净化，以甲醇-异丙醇（1:1）进行洗脱，采用Waters ACQUITY UPLC BEH C18（100 mm×2.1 mm，1.8 μm）色谱柱，以乙腈-0.1%甲酸溶液（含0.5 mmol·L-1乙酸铵）为流动相进行分离。在电喷雾离子源（ESI）正负离子模式下电离，多反应监测（MRM）模式下测定。

结果：

17个化合物在相应的测定范围内呈良好的线性关系，相关系数（r）均大于0.99，该方法的检测限（LOD）为0.01~13.32 μg·g-1，定量限（LOQ）为0.02~26.64 μg·g-1。在3种加样水平下，回收率范围为77.7%~108.5%，RSD（n=6）为0.62%~4.7%。应用本方法对35批次医用冷敷贴样品进行分析检测，结果8批次检出双氯芬酸钠，每片检出量为0.09~27.55 mg。

结论：

该方法简便，准确度高，灵敏度好，可用于医用冷敷贴中非法添加化学药物的检测，为检测医用冷敷贴中的非法添加提供了技术支持。

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Tel：18667178751；E-mail：

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Tel：18667178751；E-mail：

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Tel：18667178751；E-mail：

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Information of 17 reference compounds

, figureFileSmall=null, figureFileBig=null, tableContent=
化合物
(compound)		CAS号
(CAS No.)		分子式
(formula)		 Mr厂家
(supplier)		批号
(batch No.)		纯度
(purity)/%
对乙酰氨基酚（paracetamol）103-90-2C8H9NO2151.16NIFDC100018-202312100.0
盐酸普鲁卡因（procaine hydrochloride）51-05-8C13H20N2O2.HCl272.77NIFDC100424-20160399.2
利多卡因（lidocaine）137-58-6C14H22N2O234.34NIFDC100342-20200699.7
苯佐卡因（benzocaine）94-09-7C9H11NO2165.19NIFDC100454-20160299.9
盐酸丁卡因（tetracaine hydrochloride）136-47-0C15H24N2O2.HCl300.83NIFDC100456-201602100.0
地塞米松（dexamethasone）50-02-2C22H29FO5392.47NIFDC100129-20190799.8
吡罗昔康（piroxicam）36322-90-4C15H13N3O4S331.35NIFDC100177-20170499.5
曲安奈德（triamcinolone acetonide）76-25-5C24H31FO6434.50NIFDC100055-20210599.3
甲氧苄啶（trimethoprim）738-70-5C14H18N4O3290.32NIFDC100031-20220799.8
醋酸曲安奈德（triamcinolone acetonide acetate）3870-07-3C26H33FO7476.54NIFDC100125-20200798.4
氯倍他索丙酸酯（clobetasol propionate）25122-46-7C25H32ClFO5466.97Dr. EhrenstorferG106951498.9
保泰松（phenylbutazone）50-33-9C19H20N2O2308.37NIFDC100481-20170299.8
阿司匹林（aspirin）50-78-2C9H8O4180.16NIFDC100113-20170699.8
布洛芬（ibuprofen）15687-27-1C13H18O2206.28NIFDC100179-20230899.8
萘普生（naproxen）22204-53-1C14H14O3230.26NIFDC100198-201706100.0
双氯芬酸钠（diclofenac sodium）15307-79-6C14H10Cl2NNaO2318.13NIFDC100334-201803100.0
吲哚美辛（indometacin）53-86-1C19H16ClNO4357.79NIFDC100258-20210599.9
), ArticleFig(id=1240954734186648031, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1240945601282633762, language=CN, label=表1, caption=

17个化合物对照品信息

, figureFileSmall=null, figureFileBig=null, tableContent=
化合物
(compound)		CAS号
(CAS No.)		分子式
(formula)		 Mr厂家
(supplier)		批号
(batch No.)		纯度
(purity)/%
对乙酰氨基酚（paracetamol）103-90-2C8H9NO2151.16NIFDC100018-202312100.0
盐酸普鲁卡因（procaine hydrochloride）51-05-8C13H20N2O2.HCl272.77NIFDC100424-20160399.2
利多卡因（lidocaine）137-58-6C14H22N2O234.34NIFDC100342-20200699.7
苯佐卡因（benzocaine）94-09-7C9H11NO2165.19NIFDC100454-20160299.9
盐酸丁卡因（tetracaine hydrochloride）136-47-0C15H24N2O2.HCl300.83NIFDC100456-201602100.0
地塞米松（dexamethasone）50-02-2C22H29FO5392.47NIFDC100129-20190799.8
吡罗昔康（piroxicam）36322-90-4C15H13N3O4S331.35NIFDC100177-20170499.5
曲安奈德（triamcinolone acetonide）76-25-5C24H31FO6434.50NIFDC100055-20210599.3
甲氧苄啶（trimethoprim）738-70-5C14H18N4O3290.32NIFDC100031-20220799.8
醋酸曲安奈德（triamcinolone acetonide acetate）3870-07-3C26H33FO7476.54NIFDC100125-20200798.4
氯倍他索丙酸酯（clobetasol propionate）25122-46-7C25H32ClFO5466.97Dr. EhrenstorferG106951498.9
保泰松（phenylbutazone）50-33-9C19H20N2O2308.37NIFDC100481-20170299.8
阿司匹林（aspirin）50-78-2C9H8O4180.16NIFDC100113-20170699.8
布洛芬（ibuprofen）15687-27-1C13H18O2206.28NIFDC100179-20230899.8
萘普生（naproxen）22204-53-1C14H14O3230.26NIFDC100198-201706100.0
双氯芬酸钠（diclofenac sodium）15307-79-6C14H10Cl2NNaO2318.13NIFDC100334-201803100.0
吲哚美辛（indometacin）53-86-1C19H16ClNO4357.79NIFDC100258-20210599.9
), ArticleFig(id=1240954734316671459, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1240945601282633762, language=EN, label=Tab.2, caption=

Ion pairs and mass spectrum parameters of 17 compounds

, figureFileSmall=null, figureFileBig=null, tableContent=
化合物
(compound)		离子模式
(mode of ion)		前体离子
(precursor ion)m/z		子离子
(product ion) m/z		Q1预四极杆电压
(Q1 Pre Bias)/V		CE/VQ3预四极杆电压
(Q3 Pre Bias)/V
对乙酰氨基酚（paracetamol）+152.30110.20*-11.0-20.0-22.0
93.20-11.0-23.0-18.0
盐酸普鲁卡因（procaine hydrochloride）+236.50100.30*-17.0-17.0-19.0
120.20-16.0-26.0-12.0
利多卡因（lidocaine）+234.5086.15*-17.0-22.0-16.0
58.20-22.0-35.0-26.0
苯佐卡因（benzocaine）+166.00138.15*-12.0-16.0-26.0
77.05-12.0-27.0-30.0
盐酸丁卡因（tetracaine hydrochloride）+265.20176.20*-13.0-18.0-21.0
72.10-13.0-25.0-30.0
地塞米松（dexamethasone）+393.00355.30*-15.0-13.0-24.0
146.95-14.0-28.0-10.0
吡罗昔康（piroxicam）+332.25121.20*-25.0-21.0-26.0
164.20-14.0-17.0-29.0
曲安奈德（triamcinolone acetonide）+435.20397.30*-16.0-16.0-15.0
339.20-10.0-16.0-25.0
甲氧苄啶（trimethoprim）+291.10230.20*-14.0-24.0-25.0
261.20-14.0-25.0-27.0
醋酸曲安奈德（triamcinolone acetonide acetate）+477.20339.10*-11.0-15.0-17.0
321.25-11.0-20.0-22.0
氯倍他索丙酸酯（clobetasol propionate）+467.00373.10*-13.0-13.0-18.0
355.25-17.0-15.0-24.0
保泰松（phenylbutazone）+309.3577.15*-13.0-55.0-29.0
160.25-12.0-21.0-11.0
阿司匹林（aspirin）-179.00137.10*21.09.022.0
93.0024.021.019.0
布洛芬（ibuprofen）-205.00161.15*16.011.014.0
131.2015.017.021.0
萘普生（naproxen）-229.00169.25*26.031.017.0
170.3025.014.019.0
双氯芬酸钠（diclofenac sodium）-294.15250.20*14.014.011.0
214.2014.020.022.0
吲哚美辛（indometacin）-356.20312.25*18.011.022.0
297.2518.019.021.0
), ArticleFig(id=1240954734408946150, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1240945601282633762, language=CN, label=表2, caption=

17个化合物离子对及相关质谱参数

, figureFileSmall=null, figureFileBig=null, tableContent=
化合物
(compound)		离子模式
(mode of ion)		前体离子
(precursor ion)m/z		子离子
(product ion) m/z		Q1预四极杆电压
(Q1 Pre Bias)/V		CE/VQ3预四极杆电压
(Q3 Pre Bias)/V
对乙酰氨基酚（paracetamol）+152.30110.20*-11.0-20.0-22.0
93.20-11.0-23.0-18.0
盐酸普鲁卡因（procaine hydrochloride）+236.50100.30*-17.0-17.0-19.0
120.20-16.0-26.0-12.0
利多卡因（lidocaine）+234.5086.15*-17.0-22.0-16.0
58.20-22.0-35.0-26.0
苯佐卡因（benzocaine）+166.00138.15*-12.0-16.0-26.0
77.05-12.0-27.0-30.0
盐酸丁卡因（tetracaine hydrochloride）+265.20176.20*-13.0-18.0-21.0
72.10-13.0-25.0-30.0
地塞米松（dexamethasone）+393.00355.30*-15.0-13.0-24.0
146.95-14.0-28.0-10.0
吡罗昔康（piroxicam）+332.25121.20*-25.0-21.0-26.0
164.20-14.0-17.0-29.0
曲安奈德（triamcinolone acetonide）+435.20397.30*-16.0-16.0-15.0
339.20-10.0-16.0-25.0
甲氧苄啶（trimethoprim）+291.10230.20*-14.0-24.0-25.0
261.20-14.0-25.0-27.0
醋酸曲安奈德（triamcinolone acetonide acetate）+477.20339.10*-11.0-15.0-17.0
321.25-11.0-20.0-22.0
氯倍他索丙酸酯（clobetasol propionate）+467.00373.10*-13.0-13.0-18.0
355.25-17.0-15.0-24.0
保泰松（phenylbutazone）+309.3577.15*-13.0-55.0-29.0
160.25-12.0-21.0-11.0
阿司匹林（aspirin）-179.00137.10*21.09.022.0
93.0024.021.019.0
布洛芬（ibuprofen）-205.00161.15*16.011.014.0
131.2015.017.021.0
萘普生（naproxen）-229.00169.25*26.031.017.0
170.3025.014.019.0
双氯芬酸钠（diclofenac sodium）-294.15250.20*14.014.011.0
214.2014.020.022.0
吲哚美辛（indometacin）-356.20312.25*18.011.022.0
297.2518.019.021.0
), ArticleFig(id=1240954734517998060, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1240945601282633762, language=EN, label=Tab.3, caption=

Regression equations，correlation coefficents（r），LODs and LOQs of 17 target compounds

, figureFileSmall=null, figureFileBig=null, tableContent=
化合物
(compound)		线性范围
(linear range)/(ng·mL-1)		线性方程
（linear equation）		 rLOD/（μg·g-1）LOQ/（μg·g-1）
对乙酰氨基酚（paracetamol）7.48~897.02 Y=3.579×103X+8.908×1040.998 30.140.29
盐酸普鲁卡因（procaine hydrochloride）4.92~590.25 Y=6.541×103X+6.428×1040.998 40.050.19
利多卡因（lidocaine）4.88~585.76 Y=5.778×103X+4.864×1040.998 30.100.19
苯佐卡因（benzocaine）31.95~3 833.60 Y=8.792×102X+4.153×1040.999 60.631.25
盐酸丁卡因（tetracaine hydrochloride）0.17~20.52 Y=1.587×105X+7.380×1040.999 80.010.02
地塞米松（dexamethasone）40.65~4 878.22 Y=5.999×102X+1.429×1040.999 80.401.59
吡罗昔康（piroxicam）4.08~489.59 Y=9.773×103X-1.396×1040.999 60.040.16
曲安奈德（triamcinolone acetonide）13.62~1 634.48 Y=1.663×103X+2.673×1040.999 50.130.53
甲氧苄啶（trimethoprim）0.80~95.52 Y=6.298×104X+6.042×1040.999 60.010.02
醋酸曲安奈德（triamcinolone acetonide acetate）23.69~2 843.37 Y=2.100×104X+8.672×1040.999 30.230.46
氯倍他索丙酸酯（clobetasol propionate）6.33~1 034.68 Y=2.482×103X+1.002×1050.996 70.170.34
保泰松（phenylbutazone）7.57~908.58 Y=5.138×103X+9.688×1040.996 80.070.15
阿司匹林（aspirin）1.21~145.10 Y=3.284×104X+6.038×1040.996 311.8423.67
布洛芬（ibuprofen）339.96~40 975.16 Y=26.11X+1.473×1040.996 813.3226.64
萘普生（naproxen）246.64~29 596.80 Y=101.9X-2.771×1040.999 72.424.83
吲哚美辛（indometacin）16.51~1 981.38 Y=1.897×103X+5.95×1040.999 20.160.32
双氯芬酸钠（diclofenac sodium）22.35~2 682.40 Y=5.172×103X-1.294×1040.999 90.220.44
), ArticleFig(id=1240954734635438575, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1240945601282633762, language=CN, label=表3, caption=

17个目标化合物的回归方程、相关系数、检出限和定量限

, figureFileSmall=null, figureFileBig=null, tableContent=
化合物
(compound)		线性范围
(linear range)/(ng·mL-1)		线性方程
（linear equation）		 rLOD/（μg·g-1）LOQ/（μg·g-1）
对乙酰氨基酚（paracetamol）7.48~897.02 Y=3.579×103X+8.908×1040.998 30.140.29
盐酸普鲁卡因（procaine hydrochloride）4.92~590.25 Y=6.541×103X+6.428×1040.998 40.050.19
利多卡因（lidocaine）4.88~585.76 Y=5.778×103X+4.864×1040.998 30.100.19
苯佐卡因（benzocaine）31.95~3 833.60 Y=8.792×102X+4.153×1040.999 60.631.25
盐酸丁卡因（tetracaine hydrochloride）0.17~20.52 Y=1.587×105X+7.380×1040.999 80.010.02
地塞米松（dexamethasone）40.65~4 878.22 Y=5.999×102X+1.429×1040.999 80.401.59
吡罗昔康（piroxicam）4.08~489.59 Y=9.773×103X-1.396×1040.999 60.040.16
曲安奈德（triamcinolone acetonide）13.62~1 634.48 Y=1.663×103X+2.673×1040.999 50.130.53
甲氧苄啶（trimethoprim）0.80~95.52 Y=6.298×104X+6.042×1040.999 60.010.02
醋酸曲安奈德（triamcinolone acetonide acetate）23.69~2 843.37 Y=2.100×104X+8.672×1040.999 30.230.46
氯倍他索丙酸酯（clobetasol propionate）6.33~1 034.68 Y=2.482×103X+1.002×1050.996 70.170.34
保泰松（phenylbutazone）7.57~908.58 Y=5.138×103X+9.688×1040.996 80.070.15
阿司匹林（aspirin）1.21~145.10 Y=3.284×104X+6.038×1040.996 311.8423.67
布洛芬（ibuprofen）339.96~40 975.16 Y=26.11X+1.473×1040.996 813.3226.64
萘普生（naproxen）246.64~29 596.80 Y=101.9X-2.771×1040.999 72.424.83
吲哚美辛（indometacin）16.51~1 981.38 Y=1.897×103X+5.95×1040.999 20.160.32
双氯芬酸钠（diclofenac sodium）22.35~2 682.40 Y=5.172×103X-1.294×1040.999 90.220.44
), ArticleFig(id=1240954734731907571, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1240945601282633762, language=EN, label=Tab.4, caption=

Spiked recoveries and RSD of 17 target compounds in medical cold compress

, figureFileSmall=null, figureFileBig=null, tableContent=
化合物
（compound）		理论浓度
(theoretical concentration)/（ng·mL-1）		平均回收率
(average recovery)/%		RSD/%
高
(high) 		中
(medium)		低
(low)		高
(high) 		中
(medium)		低
(low)		高
(high) 		中
(medium)		低
(low)
对乙酰氨基酚（paracetamol）598.016149.50429.90198.296.093.61.3 2.6 3.3
盐酸普鲁卡因（procaine hydrochloride）393.49998.37519.67584.681.979.92.0 2.4 2.5
利多卡因（lidocaine）390.50597.62619.52595.990.086.12.7 2.8 3.9
苯佐卡因（benzocaine）2 555.735638.934127.78794.592.290.21.6 2.3 4.7
盐酸丁卡因（tetracaine hydrochloride）13.6833.4210.68496.589.286.42.5 2.5 1.5
地塞米松（dexamethasone）3 252.149813.037162.608101.593.091.72.5 3.0 3.9
吡罗昔康（piroxicam）326.39381.59816.32092.384.690.21.9 1.7 3.2
曲安奈德(triamcinolone acetonide)1 089.656 272.41454.483103.998.797.40.62 2.9 4.6
甲氧苄啶（trimethoprim）63.68015.9203.18486.381.477.72.2 1.7 1.8
醋酸曲安奈德（triamcinolone acetonide acetate）1 895.578473.89494.77992.684.587.92.8 2.7 3.1
氯倍他索丙酸酯（clobetasol propionate）689.787172.44734.48997.0108.1100.81.9 2.3 4.2
保泰松（phenylbutazone）605.720151.43030.28688.785.790.72.4 1.9 2.3
阿司匹林（aspirin）96 734.14424 183.5364 836.707101.1106.2101.21.8 2.2 1.9
布洛芬（ibuprofen）27 196.786 799.1941 359.839108.5100.296.21.9 1.6 4.6
萘普生（naproxen）19 731.24 932.8986.56105.194.9101.61.7 2.9 1.5
吲哚美辛（indometacin）1 320.918330.22966.04692.792.2101.72.5 2.9 4.1
双氯芬酸钠（diclofenac sodium）1 788.267447.06789.413103.996.3100.71.1 1.4 1.1
), ArticleFig(id=1240954734895485430, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1240945601282633762, language=CN, label=表4, caption=

17个目标化合物在医用冷敷贴中的回收率及RSD（n=6）

, figureFileSmall=null, figureFileBig=null, tableContent=
化合物
（compound）		理论浓度
(theoretical concentration)/（ng·mL-1）		平均回收率
(average recovery)/%		RSD/%
高
(high) 		中
(medium)		低
(low)		高
(high) 		中
(medium)		低
(low)		高
(high) 		中
(medium)		低
(low)
对乙酰氨基酚（paracetamol）598.016149.50429.90198.296.093.61.3 2.6 3.3
盐酸普鲁卡因（procaine hydrochloride）393.49998.37519.67584.681.979.92.0 2.4 2.5
利多卡因（lidocaine）390.50597.62619.52595.990.086.12.7 2.8 3.9
苯佐卡因（benzocaine）2 555.735638.934127.78794.592.290.21.6 2.3 4.7
盐酸丁卡因（tetracaine hydrochloride）13.6833.4210.68496.589.286.42.5 2.5 1.5
地塞米松（dexamethasone）3 252.149813.037162.608101.593.091.72.5 3.0 3.9
吡罗昔康（piroxicam）326.39381.59816.32092.384.690.21.9 1.7 3.2
曲安奈德(triamcinolone acetonide)1 089.656 272.41454.483103.998.797.40.62 2.9 4.6
甲氧苄啶（trimethoprim）63.68015.9203.18486.381.477.72.2 1.7 1.8
醋酸曲安奈德（triamcinolone acetonide acetate）1 895.578473.89494.77992.684.587.92.8 2.7 3.1
氯倍他索丙酸酯（clobetasol propionate）689.787172.44734.48997.0108.1100.81.9 2.3 4.2
保泰松（phenylbutazone）605.720151.43030.28688.785.790.72.4 1.9 2.3
阿司匹林（aspirin）96 734.14424 183.5364 836.707101.1106.2101.21.8 2.2 1.9
布洛芬（ibuprofen）27 196.786 799.1941 359.839108.5100.296.21.9 1.6 4.6
萘普生（naproxen）19 731.24 932.8986.56105.194.9101.61.7 2.9 1.5
吲哚美辛（indometacin）1 320.918330.22966.04692.792.2101.72.5 2.9 4.1
双氯芬酸钠（diclofenac sodium）1 788.267447.06789.413103.996.3100.71.1 1.4 1.1
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固相萃取-超高效液相色谱-串联质谱法同时测定医用冷敷贴中17个添加化学物质*
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彭彦 1 , 杨直 1 , 成立 2 , 林丽琴 1 , 陈刘伟 1
药物分析杂志 | 安全监测 2024,44(11): 1983-1991
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药物分析杂志 | 安全监测 2024, 44(11): 1983-1991
固相萃取-超高效液相色谱-串联质谱法同时测定医用冷敷贴中17个添加化学物质*
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彭彦1 , 杨直1, 成立2, 林丽琴1, 陈刘伟1
作者信息
  • 1.杭州市食品药品检验科学研究院，杭州 310022
  • 2.湖北省药品监督管理局药品审评检查中心，武汉 430077

    • Tel：18667178751；E-mail：

Determination of 17 chemicals in medical cold compress by solid phase extraction-ultra performance liquid chromatography-tandem mass spectrometry*
Yan PENG1 , Zhi YANG1, Li CHENG2, Li-qin LIN1, Liu-wei CHEN1
Affiliations
  • 1.Hangzhou Institute for Food and Drug Control，Hangzhou 310022，China
  • 2.Evaluation and Inspection Center of Hubei Medical Products Administration，Wuhan 430077，China
出版时间: 2024-12-01 doi: 10.16155/j.0254-1793.2024-0212
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摘要
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目的：

采用固相萃取-超高效液相色谱-串联质谱（SPE-UPLC-MS/MS）法建立医用冷敷贴中17个化学物质的检测方法，该法适用于常见医用冷敷贴基质。

方法：

重点考察了固相萃取柱类型、洗脱溶剂种类及溶剂用量。样品用甲醇超声提取后，与水以1:9的比例混合，经Agilent Bond Elut HLB固相萃取柱净化，以甲醇-异丙醇（1:1）进行洗脱，采用Waters ACQUITY UPLC BEH C18（100 mm×2.1 mm，1.8 μm）色谱柱，以乙腈-0.1%甲酸溶液（含0.5 mmol·L-1乙酸铵）为流动相进行分离。在电喷雾离子源（ESI）正负离子模式下电离，多反应监测（MRM）模式下测定。

结果：

17个化合物在相应的测定范围内呈良好的线性关系，相关系数（r）均大于0.99，该方法的检测限（LOD）为0.01~13.32 μg·g-1，定量限（LOQ）为0.02~26.64 μg·g-1。在3种加样水平下，回收率范围为77.7%~108.5%，RSD（n=6）为0.62%~4.7%。应用本方法对35批次医用冷敷贴样品进行分析检测，结果8批次检出双氯芬酸钠，每片检出量为0.09~27.55 mg。

结论：

该方法简便，准确度高，灵敏度好，可用于医用冷敷贴中非法添加化学药物的检测，为检测医用冷敷贴中的非法添加提供了技术支持。

超高效液相色谱-串联质谱（UPLC-MS/MS）  /  固相萃取（SPE）  /  糖皮质激素  /  解热镇痛  /  麻醉剂  /  医用冷敷贴
Objective:

To establish a method for the determination of 17 kinds of anti-allergic compounds in medical cold compress by solid phase extraction-ultra performance liquid chromatography-tandem mass spectrometry（SPE-UPLC-MS/MS）. Common medical cold compress matrices were suitable for this new method.

Methods:

Three pretreatment methods were compared：prime pass-through column purification，SPE purification including HLB and MCX solid-phase extraction. The pretreatment was optimized by focusing on the kind of solid-phase extraction，the type and volume of elution solvents. The analytes were extracted using methanol and then mixed with water at the ratio of 1:9 in volume，purified with an Agilent Bond Elut HLB column. Waters ACQUITY UPLC BEH C18（100 mm×2.1 mm，1.8 μm）column was used with 0.1% formic acid(with 0.5 mmol·L-1 ammonium acetate)-acetonitrile as mobile phases by gradient elution for separating 17 compounds. Detection was carried out by positive and negative electrospray ionization（ESI）mass spectrometer in multiple reaction monitor (MRM) mode.

Results:

All the 17 kinds of compounds showed good linearity in their reasonable ranges（r）>0.99，the limit of detection （LOD） and quantification（LOQ）were 0.01-13.32 μg·g-1 and 0.02-26.64 μg·g-1，respectively. The average recoveries of at three spiked levels were in range of 77.7%-108.5%，with RSDs （n=6） were 0.62%-4.7%. The content range of diclofenac sodium in positive samples was 0.09-27.55 mg per tablet.

Conclusion:

The method is simple，sensitive and reliable，and is suitable for determination of 17 illegally add chemical drugs in medical cold compress. Moreover，the method might provide technological support for the detection of illegal additions in medical cold compress.

ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS)  /  solid phase extraction (SPE)  /  glucocorticoids  /  antipyretic and analgesic  /  anesthetic  /  medical cold compress
彭彦, 杨直, 成立, 林丽琴, 陈刘伟. 固相萃取-超高效液相色谱-串联质谱法同时测定医用冷敷贴中17个添加化学物质*. 药物分析杂志, 2024 , 44 (11) : 1983 -1991 . DOI: 10.16155/j.0254-1793.2024-0212
Yan PENG, Zhi YANG, Li CHENG, Li-qin LIN, Liu-wei CHEN. Determination of 17 chemicals in medical cold compress by solid phase extraction-ultra performance liquid chromatography-tandem mass spectrometry*[J]. Chinese Journal of Pharmaceutical Analysis, 2024 , 44 (11) : 1983 -1991 . DOI: 10.16155/j.0254-1793.2024-0212
正文
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按《医疗器械分类目录》，医用冷敷贴的定义为“通常由降温物质和各种形式的外套及固定器具组成”，其中降温物质不应含有发挥药理学、免疫学或者代谢作用的成分。但在实际应用中，医用冷敷贴存在成分和备案不一，虚假宣传的情况，易被企业套用成械字号面膜或中药贴膏等非医疗器械产品，对外宣称械字号面膜主要用于保湿补水，中药贴膏主要用于缓解疼痛。前期市场抽验发现，某些商家为牟利，擅自非法添加糖皮质激素类、麻醉类、解热镇痛类成分，以迅速达到美白、消炎及缓解疼痛的功效[1]。我国[2]、东盟[3]等国家及地区均在法规中将激素、麻醉类明确列为禁用组分。
目前国内外关于糖皮质激素的检测多集中在化妆品，方法以液相色谱-串联质谱法[4-12]为主，其选择性好，灵敏度高，特异性强，能够快速准确测定化合物，已成为化妆品标准采用的主流分析仪器。解热镇痛类的研究多集中在保健食品及中成药中，方法主要有液相色谱-高分辨质谱法[13]、表面增强拉曼光谱法[14]、高效液相色谱法[15]、液相色谱-串联质谱法[16-17]等。针对非法添加糖皮质激素、局部麻醉剂、解热镇痛类化合物的情况，国家出台了一系列标准，但医疗器械不适用于上述标准。由于医疗器械样品基质主要为有黏附力的膏体层及表面活性剂等，需选择适宜的前处理方法减少基质的干扰，故针对医用冷敷贴非法添加亟待开展相关研究。
本研究采用固相萃取技术，运用超高效液相色谱串联三重四极杆质谱法，对医用冷敷贴中非法添加的糖皮质激素类、麻醉类及解热镇痛类等17个化学成分建立了一种快速、高效的定量检测方法，为医用冷敷贴的监管提供了技术支持。
1　仪器与试药
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超高效液相串联三重四极杆质谱仪配有DGU-20A液相色谱系统和8050 MS/MS质谱系统（岛津公司）；Milli-Q超纯水器（Millipore公司）；KQ5200V超声波清洗器（功率200 W，频率40 kHz，昆山市超声仪器有限公司）；Mettler XS205DU电子天平（精度0.01 mg，Mettler Toledo公司）。
乙腈、甲醇（色谱级，Merck公司），乙酸铵（分析纯，TCI公司），甲酸（质谱级，ACS公司）；Waters Prime HLB固相萃取柱和Waters Oasis MCX混合型阳离子交换固相萃取柱，规格均为3 cc/60 mg，沃特世科技有限公司；Bond Elut HLB固相萃取柱，规格3 cc/60 mg，安捷伦科技有限公司。35批次样品，其中28批购于线上，7批购于线下；对照品信息见表1。
2　方法与结果
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2.1　溶液制备
2.1.1　混合对照品储备液
精密称取各化合物对照品适量，用甲醇溶解并稀释成终质量浓度为0.4 mg·mL-1（吡罗昔康、对乙酰氨基酚、氯倍他索丙酸酯、盐酸普鲁卡因、利多卡因、盐酸丁卡因、甲氧苄啶）、2.0 mg·mL-1（地塞米松、曲安奈德、萘普生、醋酸曲安奈德、双氯芬酸钠、吲哚美辛、保泰松、布洛芬）、4.0 mg·mL-1（苯佐卡因）、6.0 mg·mL-1（阿司匹林）的单一成分对照品储备液，置-4 ℃冰箱中保存备用。
混合对照品储备液（临用现配）：精密量取各单一成分对照品储备液适量，用甲醇稀释成每1 mL含阿司匹林2.4 mg，含布洛芬、萘普生约500 μg，含地塞米松、苯佐卡因约80 μg，含吲哚美辛、曲安奈德醋酸酯、双氯芬酸钠约40 μg，含曲安奈德、氯倍他索丙酸酯、保泰松约20 μg，含吡罗昔康、对乙酰氨基酚、盐酸普鲁卡因、利多卡因约10 μg，含甲氧苄啶约1.6 μg，含盐酸丁卡因约0.3 μg的混合溶液，即得。
2.1.2　供试品溶液
取样品1片，以纱布覆盖黏性层，用剪刀剪成约1×1 cm2的大小后，置20 mL量瓶中，加入甲醇10 mL，超声（200 W，40 kHz）提取处理20 min，放冷至室温后，用甲醇定容至刻度，摇匀，0.45 μm滤膜过滤；精密量取续滤液1.0 mL，加水9.0 mL混匀，转移至Bond Elut HLB小柱中，待溶液完全通过后，用水2 mL淋洗，抽至近干，用甲醇-异丙醇（1:1）4 mL洗脱，抽干，收集洗脱液，经0.22 μm滤膜过滤，取续滤液，即得。
2.2　仪器条件
2.2.1　色谱条件
采用Waters BEH C18（100 mm×2.1 mm，1.8 μm）色谱柱，以乙腈(A)-0.1%甲酸溶液（含0.5 mmol·L-1乙酸铵）(B)为流动相，梯度洗脱（0~1 min，70%B；1~2 min，70%B→60%B；2~3 min，60%B→5%B；3~8 min，5%B；8~9 min，5%B→70%B；9~12 min，70%B），流速0.4 mL·min-1，柱温40 ℃，进样量1 μL。
2.2.2　质谱条件
采用电喷雾离子源（ESI），正离子或负离子模式，多反应监测（MRM）模式，离子源温度250 ℃，毛细管电压3.5 kV，干燥气流量10 L·min-1，鞘气温度250 ℃，雾化气流速10 L·min-1。17种化合物的质谱参数见表2。
分别取“2.1”项下混合对照品溶液和供试品溶液，注入液相色谱质谱联用仪进行测定，记录色谱图（图1）。
2.3　方法学考察
2.3.1　线性关系考察
精密量取“2.1.1”项下混合对照品储备液适量，用甲醇-异丙醇（1:1）稀释，配制8个浓度点的混合对照品溶液，分别按“2.2”项下色谱-质谱条件进行分析，以被测化合物定量离子对的峰面积Y为纵坐标，对照品质量浓度X（ng·mL-1）为横坐标进行线性回归拟合，得到线性方程和相关系数（r）。结果（表3）表明，17个化合物在各自线性范围内线性关系良好。
2.3.2　检测限与定量限
精密量取混合对照品溶液适量，加入阴性样品（批号220420，陕西佰傲再生医学有限公司）中，照“2.1.2”项下方法配制溶液，按“2.2”项下色谱-质谱条件进行分析，以信噪比为3和10时的含量作为17个化合物的检测限（LOD）和定量限（LOQ），结果见表3。
2.3.3　回收率试验
取“2.3.2”项下的阴性样品，分别添加低、中、高3个浓度水平的混合对照品溶液，每个浓度水平按照“2.1.2”项下方法平行制备6份供试溶液，进样测定，计算17个化合物回收率和RSD，结果见表4。
2.3.4　精密度试验
取“2.3.3”项下3个浓度点的加标溶液，每个浓度水平平行测定6次，测得17个化合物峰面积RSD为0.60%~4.6%（n=6），具体结果见表4。
2.3.5　稳定性试验
照“2.3.3”项下方法制备3个浓度点的阳性供试品溶液，分别于室温下避光放置0、2、4、6、8、10、12 h进行测定，记录峰面积，结果17个化合物12 h内峰面积的RSD均小于10%（n=6），表明供试品溶液在室温下12 h内稳定性良好。
2.4　样品测定分析
基于上述建立的方法，对收集到的线上、线下共计35批次的样品进行筛查，每份样品平行称取2份，按“2.1.2”项下方法进行处理，采用所建立的超高效液相色谱-串联质谱联用法进行定量检测，结果表明8批次样品中检出双氯芬酸钠，每片检出量由小到大依次为0.09、1.56、4.36、5.18、5.24、6.35、7.41、27.55 mg。
3　分析与讨论
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3.1　提取方法的选择
膏药贴因其一面为具有黏附力的膏体层，不易剪碎导致很难取样均匀。实验采用纱布覆盖膏药贴的黏性层，再用剪刀将其剪成约1×1 cm2的大小后，整片用提取剂浸泡提取，纱布为松散的开放式编制，不影响膏药与提取剂的接触。甲醇为较常见的提取溶剂，针对各化合物性质以及前期对照品溶液配制情况发现，17个待测化合物在甲醇中的溶解性均较好。实验也考察了不同提取方法（超声、涡旋提取）及不同超声时间（5、10、15、20、25、30 min）对17个化合物回收率的影响，发现随着提取时间的延长，提取得到的添加物组分呈现上升后平缓的趋势，即在提取时间为20 min后，延长提取时间对提取效果的增效并不明显。
3.2　流动相的选择
流动相种类、pH、比例等均会影响化合物的峰形、保留时间以及响应。本实验比较了甲醇、乙腈作为有机相，水、氨溶液、甲酸及乙酸溶液、挥发性缓冲盐（乙酸铵溶液）作为水相时各化合物的峰形以及响应情况。
结合前期质谱条件优化的情况，发现17个化合物共有2种电离模式，故优先选择了水为水相进行测试，比较了乙腈及甲醇作为有机相时各峰的峰形，结果发现不管是采用乙腈还是甲醇，吡罗昔康、丁卡因、利多卡因等均出现了严重的拖尾、分叉情况，负离子模式的阿司匹林、布洛芬响应低，且甲醇-水系统压力高。为提高负离子响应，测试了以0.1%氨溶液作为水相，甲醇、乙腈分别作为有机相时各化合物的峰形，结果发现2个系统部分组分均出现了裂峰的情况。采用0.1%甲酸溶液为水相，负离子模式的化合物响应降低，但以乙腈作为有机相时，各峰的峰形较上述其他各条件稍有改善，少数化合物有拖尾现象，而在水相中加入少量乙酸铵能消除这种拖尾现象，分析其原因可能是甲酸与乙酸铵形成缓冲体系，改善组分的峰形。进一步实验发现，乙酸铵的浓度过大反而会降低部分目标化合物的离子化效率，故结合上述测试的情况，在兼顾17个化合物峰形以及灵敏度的情况下，选用乙腈-0.1%甲酸溶液（含0.5 mmol·L-1乙酸铵）为流动相。
3.3　固相萃取小柱的选择
样品基质可能会影响目标化合物的响应，造成目标化合物响应增强或抑制的情况，为降低基质效应的影响，本实验采用了固相萃取的方式对样品前处理方法进行优化，在减少样品基质干扰的同时达到富集的目的。结合化合物结构特征及pKa值，选择了符合多数化合物可能适合的固相萃取柱（HLB固相萃取柱、MCX混合型阳离子交换固相萃取柱）。MCX混合型阳离子交换固相萃取柱兼有离子吸附和反相吸附混合作用力，适用于pKa值为2~10的化合物；Agilent Bond Elut HLB萃取柱由亲水亲脂平衡材料聚合而成，对极性和非极性的化合物均有一定的保留。不管选择哪种类型的固相萃取柱，吲哚美辛的回收率均低于30%，收集前处理过程中每一步的液体均未检出吲哚美辛，考虑吲哚美辛未能被洗脱完全，而其余组分采用HLB一步法或三步法，回收率结果差异不大；采用MCX固相萃取柱时，对乙酰氨基酚的回收率仅为36.0%，其余15个化合物的回收率在77.2%~157.0%，见图2。考虑HLB一步法可优化的参数少，HLB三步法的初筛结果优于MCX固相萃取柱，本实验进一步比较了不同类型及不同体积洗脱液对各化合物回收率的影响，见图3。结果发现，不管甲醇还是乙腈，2 mL洗脱体积无法将化合物完全洗脱，采用甲醇-异丙醇（2:1）4 mL洗脱时，阿司匹林出现了裂峰的情况，结合回收率的结果及各峰峰形，最终选择甲醇-异丙醇（1:1）4 mL进行洗脱，所有化合物的回收率均在83%~112%。
4　结论
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本研究建立了UPLC-MS/MS测定医用冷敷贴中17个化合物的检测方法，该方法具有快速、高效、准确的特点，仪器可在12 min内能完成17个非法添加类化合物的筛查和定量检测，大大提高了工作效率，可在实际应用中对样品进行快速筛查，为医用冷敷贴风险监测提供有力的技术支撑，并为其他医疗器械非法添加的检测提供了解决方案。
基金
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  • *浙江省食品药品监管系统科技计划项目(2021027)
文献
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2024年第44卷第11期
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doi: 10.16155/j.0254-1793.2024-0212
  • 接收时间：2024-03-30
  • 首发时间：2026-03-18
  • 出版时间：2024-12-01
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  • 收稿日期：2024-03-30
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*浙江省食品药品监管系统科技计划项目(2021027)
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    1.杭州市食品药品检验科学研究院，杭州 310022
    2.湖北省药品监督管理局药品审评检查中心，武汉 430077
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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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