We developed a selective bioanalytical RP-HPLC method for estimation of vitamin E from dried blood spot (DBS) sample, a potential technique which can be used for population-based epidemiological studies. Vitamin E was extracted from DBS by using liquid-liquid extraction technique with methanol (100% v/v) as reconstituting solvent for the residue. Alpha tocopheryl acetate was used as internal standard. Samples were analyzed directly on HPLC with C18 (250 × 4.6 mm × 5 μm) Phenomenex column. The mobile phase used was methanol to water (99:1% v/v) at a flow rate of 1.4 mL/min. The detector wavelength used was 292 nm. The retention time observed for vitamin E and internal standard was 10.225 ± 0.00075 min and 13.580 ± 0.00075 min respectively. The vitamin E calibration curve was found to be linear over the range of 0.625 to 60 μg/mL. The limit of quantification for vitamin E was found to be 0.1 μg/mL. Accuracy of the developed method was found to be 103.179%, 101.625%, and 100.174% with percentage of coefficient of variation of 0.0161, 0.0215, and 0.2790 for HQC, MQC, and LQC samples respectively which were within USFDA acceptance limit of ± 15 to ± 20%.The intraday and interday precision expressed as coefficient of variation were 0.0191–0.0841% and 0.0074–0.0252% respectively. The method represents a simple, rapid, specific, accurate, and precise method for estimation of vitamin E in human blood using DBS technique. The developed method can be further evaluated with respect to effect of matrix variability before it can be used in clinical setting.

中文翻译：

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色谱法从干血斑样品中估算维生素E

我们开发了一种选择性的生物分析RP-HPLC方法，用于从干血斑（DBS）样品中估算维生素E，这是一种可用于基于人群的流行病学研究的潜在技术。采用液-液萃取技术，以甲醇（100％v / v）为残留物的重构溶剂，从星展银行中提取维生素E。醋酸α生育酚酯用作内标。使用C18（250×4.6 mm×5μm）Phenomenex色谱柱在HPLC上直接分析样品。所用的流动相为甲醇至水（99：1％v / v），流速为1.4 mL / min。使用的检测器波长为292 nm。维生素E和内标物的保留时间分别为10.225±0.00075分钟和13.580±0.00075分钟。发现维生素E校准曲线在0.625至60μg/ mL的范围内呈线性。维生素E的定量限为0.1μg/ mL。发现该方法的准确度为103.179％，101.625％和100.174％，HQC，MQC和LQC样品的变异系数分别为0.0161、0.0215和0.2790，在USFDA接受极限的±15至±20％。用变异系数表示的日内和日间精度分别为0.0191–0.0841％和0.0074–0.0252％。该方法代表使用DBS技术估算人血中维生素E的简单，快速，特异，准确和精确的方法。在将其用于临床之前，可以针对矩阵变异性的效果进一步评估所开发的方法。HQC，MQC和LQC样品的174％的变异系数分别为0.0161、0.0215和0.2790，在USFDA接受极限的±15至±20％之内。日内和日间精度表示为0.0191分别为–0.0841％和0.0074–0.0252％。该方法代表使用DBS技术估算人血中维生素E的简单，快速，特异，准确和精确的方法。在将其用于临床之前，可以针对矩阵变异性的效果进一步评估所开发的方法。HQC，MQC和LQC样品的174％的变异系数分别为0.0161、0.0215和0.2790，在USFDA接受极限的±15至±20％之内。日内和日间精度表示为0.0191分别为–0.0841％和0.0074–0.0252％。该方法代表使用DBS技术估算人血中维生素E的简单，快速，特异，准确和精确的方法。在将其用于临床之前，可以针对矩阵变异性的效果进一步评估所开发的方法。分别为0074–0.0252％。该方法代表使用DBS技术估算人血中维生素E的简单，快速，特异，准确和精确的方法。在将其用于临床之前，可以针对矩阵变异性的效果进一步评估所开发的方法。分别为0074–0.0252％。该方法代表一种使用DBS技术估算人血中维生素E的简单，快速，特异，准确和精确的方法。在将其用于临床之前，可以针对矩阵变异性的效果进一步评估所开发的方法。