Preparation of Reference Materials for Carbon and Nitrogen Isotopes of Organic Materials
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摘要: 实验室和研究人员所使用的碳、氮同位素标准物质一般由国际原子能机构（IAEA）获得，然而近年来，随着碳氮同位素在实验室质量监控、方法评价、仪器校准等方面的广泛应用，市场需求量不断增加，IAEA研制的碳、氮同位素标准物质的种类与数量逐渐不能满足科学研究快速发展的需求。我国急需研制适应当今分析技术水平的有机质碳氮同位素国家标准物质用以进行质量监控、方法评价、仪器校准。为保证量值传递精度，本文研制了4个有机化学物质的碳氮稳定同位素标准物质，其中3个为尿素样品，1个为L-谷氨酸。经检验4种标准物质的均匀性通过F值检验，标准物质的δ13C和δ15N值经过一年的稳定性检验，特征量值变化在测量方法允许的不确定度范围内，由此判定δ13C和δ15N值稳定性良好。由包括研制单位实验室在内的12家实验室协同定值，采用高温燃烧-气体同位素质谱法测定了δ13C和δ15N值，系列标准物质δ13C和δ15N认定值区间呈梯度分布，δ13C值为-40‰~0‰，δ15N值为-10‰~30‰，涵盖了我国天然样品中有机质碳氮稳定同位素组成范围；研制的系列标准物质δ13C的定值扩展不确定度不大于0.08‰，δ15N的定值扩展不确定度不大于0.09‰，定值水平与国际标准物质相当。该系列标准物质已被国家质检总局批准为国家一级标准物质，批准号为GBW04494~GBW04497。可被用于地质、生态、环境等多种样品δ13C和δ15N比值测定时的分析监控、仪器校准、方法评价、质量保证和质量监控。Abstract:
OBJECTIVES The carbon and nitrogen isotope reference materials used by laboratories and researchers are generally obtained by the International Atomic Energy Agency (IAEA). However, in recent years, with the widespread use of carbon and nitrogen isotopes in laboratory quality control, method evaluation, and instrument calibration, the market demand continues to increase, and the types and quantities of carbon and nitrogen isotope reference materials developed by IAEA are gradually unable to meet the rapid development of scientific research. The development of national standard materials for carbon and nitrogen isotopes of organic matter that are suitable for the current analytical technology level for quality control, method evaluation, and instrument calibration are urgently needed in China. OBJECTIVES To prepare carbon and nitrogen isotope reference materials for organic matter. METHODS In order to ensure the accuracy of the transmission of the value, 4 stable carbon and nitrogen isotope reference materials of organic chemical materials were developed, of which 3 were urea samples and 1 was L-glutamic acid. RESULTS According to the homogeneity test results, no statistically significant heterogeneity was found based on F test. The verified results by variance analysis for four samples showed very good homogeneity for the series. Nearly one-year stability analysis of δ13C and δ15N values confirmed that no statistically significant instability was found. The certified values of the reference materials were determined by an interlaboratory comparison of 12 participating laboratories. Carbon and nitrogen stable isotope ratios were measured using an elemental analyzer coupled with an isotope ratio mass spectrometer. The δ13C and δ15N values of the series of reference materials had a gradient distribution. The δ13C values ranged from -40‰ to 0‰, and the δ15N value from -10‰ to 30‰, covering the range of organic carbon and nitrogen stable isotope composition in natural samples in China. The extended uncertainty of the series of standard materials was not greater than 0.08‰ for δ13C and not greater than 0.09‰ for δ15N, and the fixed value level was equivalent to that of the international standard materials. This series of reference materials had been approved by the General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China as the national first-level reference materials (GBW04494-GBW04497). Conclusion The series of reference materials can be used for analysis and monitoring, instrument calibration, method evaluation, quality assurance and quality monitoring in the determination of δ13C and δ15N ratios of various samples in geology, ecology and environment. -
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表 1 碳氮同位素标准物质候选物设计
Table 1. Design of candidates for carbon and nitrogen isotopes reference materials
候选物
编号δ13CVPDB值
区间(‰)δ15NAir-N2值
区间(‰)样品制备 CN-1
(尿素1#)-46.0~-45.0 -0.5~0.5 购置商用分析纯试剂。
样品经真空干燥、研磨，过筛后分装CN-2
(尿素2#)-27.0~-26.0 33~34 人工配制。加入高富集15N(98.5%)、13C(99.%)尿素标记物，通过重量法配制。配制过程包括混合、搅拌溶解、冷冻干燥、研磨，过筛后分装 CN-3
(尿素3#)1.5~2.5 17~18 人工配制。加入高富集15N(98.5%)、13C(99.%)尿素标记物，通过重量法配制。配制过程包括混合、搅拌溶解、冷冻干燥、研磨，过筛后分装 CN-4
(L-谷氨酸)-11.5~-11.0 -8.0~-7.0 购置商用分析纯试剂。
样品经真空干燥、研磨，过筛后分装
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表 2 候选物δ13C和δ15N值的均匀性检验方差分析结果
Table 2. Variance analysis for δ13C and δ15N values homogeneity inspection of reference materials
候选物
编号检验项目 单元间平方和
(SSamong)单元内平方和
(SSwith)单元间自由度
(νamong)单元内自由度
(νwith)检验统计值
(Fcal)标准偏差
(‰)自由度临界值
(Fcrit)CN-1 δ13C
δ15N0.1557
0.16090.1418
0.152924
2425
251.14
1.100.06
0.061.97 CN-2 δ13C
δ15N0.1107
0.17020.1115
0.170524
2425
251.03
1.040.05
0.061.97 CN-3 δ13C
δ15N0.1797
0.13840.1629
0.124224
2425
251.15
1.160.06
0.051.97 CN-4 δ13C
δ15N0.1116
0.14340.0871
0.144324
2425
251.33
1.040.05
0.051.97 注：方差分析表均依据《标准物质定值的通用原则及统计学原理》(JJF 1343—2012)进行统计。
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表 3 候选物δ13C和δ15N值的稳定性检验结果
Table 3. Results of for δ13C and δ15N values stability inspection of standard materials
样品编号 特征量值 T1(0个月) T2(3个月) T3(6个月) T4(12个月) T5(19个月) CN-1 δ13C(‰) -45.56 (n=4) -45.61 (n=4) -45.54 (n=4) -45.60 (n=4) -45.58 (n=4) δ15N(‰) -0.26 (n=4) -0.22 (n=4) -0.27 (n=4) -0.25 (n=4) -0.28 (n=4) CN-2 δ13C(‰) -26.55 (n=4) -26.52 (n=4) -26.55 (n=4) -26.57 (n=4) -26.54 (n=4) δ15N(‰) 33.79 (n=4) 33.77 (n=4) 33.87 (n=4) 33.85 (n=4) 33.88 (n=4) CN-3 δ13 2.15 (n= 4) 2.20 (n=4) 2.18 (n=4) 2.14 (n=4) 2.15 (n=4) δ15N 17.76 (n=4) 17.78 (n=4) 17.81 (n=4) 17.74 (n=4) 17.77 (n=4) CN-4 δ13C(‰) -11.09 (n=4) -11.06 (n=4) -11.11 (n=4) -11.08 (n=4) -11.12 (n=4) δ13C(‰) -11.09 (n=4) -11.06 (n=4) -11.11 (n=4) -11.08 (n=4) -11.12 (n=4)
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表 4 各实验室δ13C值的分析方法和分析数据
Table 4. Multiple measurement data of δ13C for the four reference materials
实验室编号 CN-1 CN-2 CN-3 CN-4 1 -45.62 -26.61 2.14 -11.09 2 -45.71 -26.55 2.25 -11.10 3 -45.52 -26.60 2.21 -11.06 4 -45.60 -26.57 2.18 -11.08 5 -45.63 -26.55 2.10 -11.09 6 -45.91 -26.62 2.10 -11.06 7 -45.60 -26.57 2.15 -11.10 8 -45.60 -26.62 2.23 -11.18 9 -45.60 -26.55 2.25 -11.11 10 -45.51 -26.60 2.01 -11.10 平均值(‰) -45.60 -26.58 2.16 -11.09 标准偏差s(‰) 0.058 0.029 0.078 0.016 标准不确定度uA 0.0183 0.0089 0.0234 0.0050
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表 5 各实验室δ15N值的分析方法和分析数据
Table 5. Multiple measurement data of δ15N for the four reference materials
实验室编号 CN-1 CN-2 CN-3 CN-4 1 -0.25 33.79 17.67 -7.52 1 -0.25 33.81 17.76 -7.56 1 -0.24 33.78 17.77 -7.52 2 -0.25 33.80 17.72 -7.49 2 -0.29 33.71 17.69 -7.48 4 -0.21 33.62 17.64 -7.41 5 -0.24 33.78 17.74 -7.51 6 -0.20 33.90 17.84 -7.53 8 -0.25 33.78 17.62 -7.56 9 -0.25 33.61 17.64 -7.50 10 -0.16 33.77 17.76 -7.50 11 -0.31 33.63 17.64 -7.58 12 -0.25 33.79 17.67 -7.52 平均值(‰) -0.24 33.75 17.71 -7.51 标准偏差s(‰) 0.039 0.088 0.068 0.053 标准不确定度uA 0.0114 0.0256 0.0192 0.0127
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表 6 定值元素δ13C和δ15N认定值及扩展不确定度
Table 6. δ13C and δ15N ceritified values for materials GBW04494 to GBW04497 and their associated expanded uncertainties (k=2)
同位素比值 认定值及扩展不确定度(‰) GBW04494 GBW04495 GBW04496 GBW04497 δ13CVPDB -45.60±0.08 -26.58±0.06 2.16±0.10 -11.09±0.09 δ15NAir-N2 -0.24±0.13 33.75±0.09 17.71±0.09 -7.51±0.06
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