伏马毒素B<sub>1</sub>纯度标准物质的研制

郭文博; 孟佳佳; 牛雪珂; 王欣艺; 聂冬霞; 赵志辉; 韩铮

doi:10.12338/j.issn.2096-9015.2023.0284

伏马毒素B₁纯度标准物质的研制

doi: 10.12338/j.issn.2096-9015.2023.0284

郭文博^{1, 2,},
孟佳佳¹,
牛雪珂^{1, 2},
王欣艺¹,
聂冬霞¹,
赵志辉¹,
韩铮^1, ,

1.
上海市农业科学院，上海 201403
2.
上海科立特农产品检测技术服务有限公司，上海 201403

基金项目: 上海市科技兴农技术创新项目（沪农科创字（2012）第3-2号）；上海市“科技创新行动计划”启明星项目（21QB1403300）。

详细信息

作者简介:
郭文博（1988-），上海市农业科学院助理研究员，研究方向：真菌毒素标准物质研制与应用，邮箱：guo1103bo@126.com

通讯作者:
韩铮（1983-），上海市农业科学院研究员，研究方向：农产品质量安全，邮箱：hanzheng@saas.sh.cn

中图分类号: TB99
计量
- 文章访问数: 131
- HTML全文浏览量: 61
- PDF下载量: 288
- 被引次数: 0
出版历程
- 收稿日期: 2023-11-17
- 录用日期: 2023-11-20
- 修回日期: 2023-11-29
- 网络出版日期: 2023-12-06
- 刊出日期: 2023-10-18

Development of a Certified Reference Material for Fumonisin B₁ Purity

GUO Wenbo^{1, 2
,},
MENG Jiajia¹,
NIU Xueke^{1, 2},
WANG Xinyi¹,
NIE Dongxia¹,
ZHAO Zhihui¹,
HAN Zheng^{1
, ,}

1.
Shanghai Academy of Agricultural Sciences, Shanghai 201403, China
2.
Shanghai Kelite Agricultural Product Testing Technology Service Co., Ltd., Shanghai 201403, China

摘要

摘要: 为更好的满足农产品质量安全检测需求，保证检测结果的准确性、可靠性和可溯源性，研制了伏马毒素B₁（FB₁）纯度标准物质。以实验室自主分离纯化获得的FB₁为候选物，通过质谱、红外光谱、核磁共振谱等对其进行定性鉴定。采用质量平衡法和定量核磁法两种不同原理的方法对候选物的纯度进行定值，其中质量平衡法中采用面积归一化法、卡尔费休法、气相色谱串联质谱法和电感耦合等离子体质谱法，分别对候选物中主成分（FB₁）、水分、挥发性杂质和非挥发性杂质含量进行测定。此外，开展了均匀性检验、稳定性考察，并对研制过程中产生的不确定度进行了系统的评定。结果表明：FB₁纯度标准物质量值为99.1%，扩展不确定度为0.3%（k=2），均匀性良好，且满足12个月的稳定性要求。研制的FB₁纯度标准物质获得国家二级标准物质证书，编号为GBW（E）100550，能够用于农产品中FB₁的定性和定量检测、方法评价以及FB₁量值溯源体系的建立。
- 计量学 /
- 伏马毒素B₁ /
- 纯度标准物质 /
- 质量平衡法 /
- 定量核磁法 /
- 不确定度评定
Abstract: To enhance the accuracy, reliability, and traceability of agricultural product quality safety testing, a certified reference material (CRM) for fumonisin B₁ (FB₁) purity was developed. The CRM candidate, obtained through laboratory purification of FB₁, was qualitatively identified using mass spectrometry, infrared spectroscopy, and nuclear magnetic resonance spectroscopy. The purity of the candidate was quantified using both mass balance and quantitative nuclear magnetic resonance methods. The mass balance method included area normalization, the Karl Fischer method, gas chromatography-tandem mass spectrometry, and inductively coupled plasma mass spectrometry to determine the main component, moisture, volatile and non-volatile impurity contents. Furthermore, homogeneity tests and stability studies were conducted, along with systematic uncertainty evaluations. The results indicated that the FB₁ CRM has a purity value of 99.1% with an extended uncertainty of 0.3% (k=2), exhibiting good homogeneity and fulfilling a 12-month stability requirement. The developed FB₁ CRM, certified as a national secondary standard material (GBW (E) 100550), is suitable for qualitative and quantitative detection, method evaluation, and establishing a traceability system for FB₁ in agricultural products
- metrology /
- fumonisin B₁ /
- purity certified reference material /
- mass balance method /
- quantitative nuclear magnetic method /
- uncertainty evaluation

HTML全文

图 1 FB₁标准物质候选物的红外光谱图

Figure 1. Infrared spectroscopy of FB₁ certified reference material candidate

下载: 全尺寸图片幻灯片

图 2 FB₁标准物质候选物的高分辨质谱图

Figure 2. Time-of-flight mass spectrometry (TOF/MS-MS) of the FB₁ certified reference material candidate

下载: 全尺寸图片幻灯片

图 3 FB₁标准物质候选物的UHPLC-CAD色谱图

Figure 3. Ultra-high performance liquid chromatography with charged aerosol detection (UHPLC-CAD) chromatograms of the FB₁ certified reference material candidate

下载: 全尺寸图片幻灯片

图 4 FB₁标准物质候选物和苯甲酸混合样品在氘代甲醇中的¹H-NMR 谱图

Figure 4. Proton nuclear magnetic resonance (¹H-NMR) spectra of mixed samples of FB₁ certified reference material candidate and benzoic acid in deuterated methanol

下载: 全尺寸图片幻灯片

表 1 FB₁标准物质候选物的质量平衡法定值结果

Table 1. Quantitative determination of the FB₁ certified reference material candidate using the mass balance method (%)

项目	结果	平均值	标准偏差
主成分纯度	99.31	99.31	0.031
	99.29
	99.32
	99.31
	99.29
	99.31
水分含量	0.143	0.139	0.003
	0.136
	0.134
	0.141
	0.143
	0.137
无机离子杂质	0.043	0.050	0.005
	0.057
	0.051
	0.044
	0.055
	0.050
纯度	99.12

下载: 导出CSV

表 2 FB₁物质候选物均匀性数据与检验结果

Table 2. Data analysis and homogeneity testing (F-test) of the FB₁ certified reference material candidate (%)

瓶号	1		2	3
1	99.35		99.34	99.31
2	99.35		99.33	99.34
3	99.29		99.29	99.32
4	99.34		99.28	99.31
5	99.32		99.27	99.31
6	99.30		99.32	99.29
7	99.26		99.32	99.34
8	99.28		99.31	99.32
9	99.29		99.32	99.33
10	99.30		99.29	99.25
11	99.34		99.29	99.31
12	99.34		99.28	99.37
13	99.34		99.29	99.31
14	99.29		99.31	99.32
15	99.32		99.31	99.28
平均值		99.31
组间方差（M_between）		0.000671
组内方差（M_within）		0.000667
F (M_between/M_within)		1.01
F_0.05（14, 30）		2.04

下载: 导出CSV

表 3 FB₁物质候选物稳定性数据与检验结果

Table 3. Analysis of short-term and long-term stability (t-test) of the FB₁ certified reference material candidate (%)

时间（天）	短期稳定性			时间（月）	长期稳定性
时间（天）	−20 ℃	4 ℃	60 ℃	时间（月）	−20 ℃
0	99.34	99.31	99.29	0	99.32
2	99.31	99.29	99.31	1	99.29
4	99.32	99.33	99.32	3	99.31
6	99.29	99.30	99.31	6	99.33
8	99.31	99.31	99.30	12	99.29
β₁	−0.0040	0.0005	0.0010	β₁	−0.0010
β₀	99.330	99.306	99.302	β₀	99.313
s	0.0151	0.0170	0.0126	s	0.0198
s(β₁)	0.0024	0.0027	0.0020	s(β₁)	0.0021
t_{0.95, 3}	3.18	3.18	3.18	t_{0.95, 3}	3.18
s(β₁)·t_0.95,3	0.0076	0.0086	0.0064	s(β₁)·t_{0.95, 3}	0.0065

下载: 导出CSV

表 4 FB₁标准物质候选物不确定度评定结果

Table 4. Evaluation of uncertainties associated with the developed FB₁ certified reference material candidate

来源	公式	不确定度/%
均匀性（u_bb）	$ {{u}}_{{{\mathrm{bb}}}}{=}\sqrt{\dfrac{{{M}}_{{{\mathrm{between}}}}{-}{{M}}_{{{\mathrm{within}}}}}{{n}}} $	0.0012
长期稳定性（u_lst）	$ {{u}}_{{{\mathrm{lts}}}}={s(}{{\beta }}_{{1}}{)}\times {t} $	0.0247
短期稳定性（u_sts）	$ {{u}}_{{{\mathrm{sts}}}}={s(}{{\beta }}_{{1}}{)}\times {t} $	0.0215
质量平衡法（u_MB）	$ {{u}}_{{{\mathrm{MB}}}}{}{=}{P}_{{{\mathrm{MB}}}}\times \sqrt{{\left[\dfrac{{u}\left({{P}}_{{0}}\right)}{{{P}}_{{0}}}\right]}^{{2}}{+}\dfrac{{{[}{u}{(}{{X}}_{{{\mathrm{W}}}}{)]}}^{{2}}{+}{{[}{u}{(}{{X}}_{{{\mathrm{NV}}}}{)]}}^{{2}}}{{{(1-}{{X}}_{{{\mathrm{W}}}}{-}{{X}}_{{{\mathrm{NV}}}}{)}}^{{2}}}} $	0.0513
定量核磁法（u_qNMR）	$ {{u}}_{{ {\mathrm{}}}}={{P}}_{{{\mathrm{FB1}}}}\sqrt{{\left[\dfrac{{u}{(}{{I}}_{{{\mathrm{FB1}}}}/{{I}}_{{{\mathrm{std}}}}{)}}{{{I}}_{{{\mathrm{FB1}}}}{/}{{I}}_{{{\mathrm{std}}}}}\right]}^{{2}}{+}{\left[\dfrac{{u}{(}{{M}}_{{{\mathrm{FB1}}}}{)}}{{{M}}_{{{\mathrm{FB1}}}}}\right]}^{{2}}{+}{\left[\dfrac{{u}{(}{{M}}_{{{\mathrm{std}}}}{)}}{{{M}}_{{{\mathrm{std}}}}}\right]}^{{2}}{+}{\left[\dfrac{{u}{(}{{m}}_{{{\mathrm{std}}}}{)}}{{{m}}_{{{\mathrm{std}}}}}\right]}^{{2}}{+}{\left[\dfrac{{u}{(}{{m}}_{{{\mathrm{FB1}}}}{)}}{{{m}}_{{{\mathrm{FB1}}}}}\right]}^{{2}}{+}{\left[\dfrac{{u}{(}{{P}}_{{{\mathrm{std}}}}{)}}{{{P}}_{{{\mathrm{std}}}}}\right]}^{{2}}} $	0.2821
定值合成不确定度（u_char）	$ {{u}}_{{{\mathrm{char}}}}{=}\dfrac{\sqrt{{{u}}_{{{\mathrm{MB}}}}^{{2}}{+}{{u}}_{{{\mathrm{qNMR}}}}^{{2}}}}{{2}} $	0.1434
合成不确定度（u_CRM）	$ {{u}}_{{{\mathrm{CRM}}}}{=}\sqrt{{{u}}_{{{\mathrm{char}}}}^{{2}}{+}{{u}}_{{{\mathrm{bb}}}}^{{2}}{+}{{u}}_{{{\mathrm{sts}}}}^{{2}}{+}{{u}}_{{{\mathrm{lst}}}}^{{2}}} $	0.1471
扩展不确定度（U）	U=k×u_CRM, k=2	0.3

下载: 导出CSV

参考文献(32)

[1]	KAMLE M, MAHATO D K, DEVI S, et al. Fumonisins: Impact on agriculture, food, and human health and their management strategies[J]. Toxins, 2019, 11(6): 328. doi: 10.3390/toxins11060328
[2]	郭志青, 张霞, 刁立功, 等. 镰刀菌及其伏马毒素的危害和防控[J]. 山东农业科学, 2022, 54(1): 157-164.
[3]	PONCE-GARCíA N, SERNA-SALDIVAR S O, GARCIA-LARA S. Fumonisins and their analogues in contaminated corn and its processed foods – a review [J]. Food Additives & Contaminants: Part A, 2018: 2183-2203.
[4]	CHEN J, WEI Z, WANG Y, et al. Fumonisin B1 : mechanisms of toxicity and biological detoxification progress in animals [J]. Food and Chemical Toxicology, 2021, 149(3): 111977.
[5]	DEEPTHI B V, SOMASHEKARAIAH R, POORNACHANDRA R K, et al. Lactobacillus plantarum MYS6 ameliorates fumonisin B1-induced hepatorenal damage in broilers[J]. Frontiers in Microbiology, 2017, 8: 2317. doi: 10.3389/fmicb.2017.02317
[6]	SORIANO J M, GONZáLEZ L, CATALá A I. Mechanism of action of sphingolipids and their metabolites in the toxicity of fumonisin B1[J]. Progress in Lipid Research, 2005, 44(6): 345-56. doi: 10.1016/j.plipres.2005.09.001
[7]	OSTRY V, MALIR F, TOMAN J, et al. Mycotoxins as human carcinogens-the IARC Monographs classification[J]. Mycotoxin research, 2017, 33: 65-73. doi: 10.1007/s12550-016-0265-7
[8]	U. S. Food and Drug Administration. guidance for industry: fumonisin levels in human foods and animal feeds, final guidance[EB/OL].https://www.fda.gov/regulatory-information/search-fda-guidance-documents/guidance-industry-fumonisin-levels-human-foods-and-animal-feeds.
[9]	The codex committee on food additives and contaminants. general standard for contaminants and toxins in food and feed : CODEX STAN 193-1995 [S]. Washington DC: FAO, 1995.
[10]	The Commission of the European Communities. setting maximum levels for certain contaminants in food stuffs : EC No 1881/2006 [S]. Brussel: official journal of the European Union, 2006.
[11]	国家质量监督检验检疫总局, 中国国家标准化管理委员会. 饲料卫生标准: GB 13078-2017[S]. 北京: 中国标准出版社, 2017.
[12]	谢刚, 王松雪. 粮油真菌毒素检测技术及标准物质研究进展[C]. 北京: 国家真菌毒素防控科技创新联盟成立大会暨第一届中国真菌毒素大会论文集, 2016.
[13]	韩铮, 郭文博, 范楷, 等. 真菌毒素检测及相关标准物质制备技术研究 [C]. 北京: 国家真菌毒素防控科技创新联盟成立大会暨第一届中国真菌毒素大会论文集, 2016.
[14]	卢晓华, 薄梦, 吴雪, 等. 标准物质领域发展现状及趋势[J]. 化学试剂, 2022, 44(10): 1403-1410.
[15]	GUO Z, LI X, LI H. Certified reference materials and metrological traceability for mycotoxin analysis [M]. Oxford University Press, 2019: 1695-707.
[16]	TANGNI E K, DEBONGNIE P, HUYBRECHTS B, et al. Towards the development of innovative multi-mycotoxin reference materials as promising metrological tool for emerging and regulated mycotoxin analyses[J]. Mycotoxin Research, 2016, 33(1): 1-10.
[17]	OLIVARES I R B, SOUZA G B, NOGUEIRA A R A, et al. Trends in developments of certified reference materials for chemical analysis-focus on food, water, soil, and sediment matrices[J]. TrAC Trends in Analytical Chemistry, 2018, 100: 53-64. doi: 10.1016/j.trac.2017.12.013
[18]	郑子繁, 刘卫晓, 金芜军, 等. 质量平衡法及其在标准物质定值中的应用进展[J]. 生物技术进展, 2020, 10(6): 623-629.
[19]	杨梦瑞, 简凌波, 王敏, 等. 盐酸沙拉沙星纯度定值方法研究及标准物质研制[J]. 农产品质量与安全, 2020(6): 35-44.
[20]	张思遥, 李晓敏, 王海峰, 等. 差示扫描量热法测定4-正辛基酚, 炔雌醇等5种化合物的纯度 [J]. 计量科学与技术, 2022, (66)7: 22-27.
[21]	韦棋, 苏福海. 甲卡西酮纯度标准物质的研制[J]. 计量科学与技术, 2020(11): 10-16.
[22]	李硕, 张楠, 刘喆, 等. α-熊果苷纯度标准物质的研制[J]. 计量科学与技术, 2022, 66(8): 7-12.
[23]	GUO W B, HAN Z, YANG J H, et al. Simultaneous preparation and characterization of three high-purity type B fumonisins from maize culture[J]. Analytical Methods, 2016, 8: 2737. doi: 10.1039/C5AY03307A
[24]	国家市场监督管理总局. 标准物质的定值及均匀性、稳定性评估: JJF 1343-2022 [S]. 北京: 中国质检出版社, 2022.
[25]	Bezuidenhout S C, Gelderblom W C A, Gorst-Allman C P, et al. Structure elucidation of the fumonisins, mycotoxins from Fusarium moniliforme[J]. Journal of the Chemical Society, Chemical Communications, 1988(11): 743-745. doi: 10.1039/c39880000743
[26]	李莉, 李硕. QuEChERS-超高效液相色谱-串联质谱法测定玉米油中伏马毒素B1, B2, B3[J]. 食品安全质量检测学报, 2020, 11(19): 7006-7011.
[27]	HAN Z, REN Y, LIU X, et al. A reliable isotope dilution method for simultaneous determination of fumonisins B1, B2 and B3 in traditional Chinese medicines by ultra‐high‐performance liquid chromatography‐tandem mass spectrometry[J]. Journal of Separation Science, 2010, 33(17-18): 2723-2733. doi: 10.1002/jssc.201000423
[28]	SZEKERES A, LORANTFY L, BENCSIK O, et al. Rapid purification method for fumonisin B1 using centrifugal partition chromatography[J]. Food Additives & Contaminants, 2013, 30(1): 147-155.
[29]	MARIA, MÅNSSON, MARIE, et al. Isolation and NMR Characterization of Fumonisin B₂ and a New Fumonisin B₆from Aspergillus niger[J]. Journal of Agricultural & Food Chemistry, 2010, 58: 949-953.
[30]	马康, 苏福海, 王海峰, 等. 有机纯度标准物质定值技术研究进展[J]. 分析测试学报, 2013, 32(7): 901-908.
[31]	WESTWOOD S, TAICHIHUANG, TINGGARRIDO, et al. Development and validation of a suite of standards for the purity assignment of organic compounds by quantitative NMR spectroscopy[J]. Metrologia, 2019, 56: 064001. doi: 10.1088/1681-7575/ab45cb
[32]	国家市场监督管理总局. 纯度标准物质定值计量技术规范有机物纯度标准物质: JJF 1855-2020 [S]. 北京: 中国质检出版社, 2020.