Development and Validation of the Ultra Performance Liquid Chromatography-Tandem Mass Spectrometer Method for Quantifica
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Development and Validation of the Ultra Performance Liquid Chromatography-Tandem Mass Spectrometer Method for Quantification of Methylenecyclopropylglycine in Litchi Fruits Using the Standard Addition Method Somya Asthana 1,2 & Kajal Karsauliya 3 & Sumita Dixit 1 & Anurag Tripathi 1 & Arvind Kumar 2 & Sheelendra Pratap Singh 3 & Mukul Das 1 Received: 16 November 2018 / Accepted: 14 May 2019 # Springer Science+Business Media, LLC, part of Springer Nature 2019
Abstract Recently, the presence of methylenecyclopropylglycine (MCPG) found in litchi seeds and ripe and unripe litchi fruit pulp proved to be a breakthrough finding as it causes hypoglycemic encephalopathy leading to death of under nourished/starved children. Earlier methods of isolation and detection of MCPG in litchi fruit were tedious and time-consuming. To reduce the complexity, we developed a better and straightforward methodology using ultra performance liquid chromatography-tandem mass spectrometer (UPLC-MS/MS) and QuEChERS technique to quantify MCPG without derivatization in litchi samples. The UPLC system included a Thermo C8 column with a mobile phase 0.1% formic acid in acetonitrile and water (gradient flow) at a flow rate of 0.3 ml/min. MCPG levels were determined using an API 4000 mass spectrometer with electrospray ionization (ESI) source. Other parameters were also determined following the quantitation by multiple reaction monitoring mode. The method was validated for selectivity, specificity, linearity, limit of quantification (LOQ), limit of detection (LOD), intra-inter day precision and accuracy, stability, and ruggedness. Calibration curve ranged from 7.8 to1000 ng/ml linearly while the LOD value for MCPG was 0.66 ng/ml. The percent accuracy and precision ranged from 92.98 to 105% and 1.78 to 8.43%, respectively. Validated method was applied for the determination of MCPG in litchi samples which provided a better and novel method for the determination of MCPG. Keywords Methylenecyclopropylglycine . Hypoglycemic encephalopathy . QuEChERS technique . Derivatization . Mass spectrometer
Introduction Acute encephalitis syndrome (AES) remains a long-standing mysterious disease in Muzzaffarpur, Bihar, even after two
* Sheelendra Pratap Singh [email protected] * Mukul Das [email protected] 1
Food Toxicology Division, Food, Drugs and Chemical Toxicology Group, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), Lucknow, India
2
School of Biotechnology, Institute of Science, Banaras Hindu University (BHU), Varanasi, U.P. 221005, India
3
Analytical Chemistry Laboratory, Regulatory Toxicology Group, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), Lucknow, U.P., India
decades due to the absence of any viral causative agent or any observable clinical diagnosis irrespective of the earlier examinations (Board 2015). The disease occurs as an annual seasonal outbreak affecting hundreds of children (below 15 years) of lower economic strata with 40–60% mortality. As hypothesized earlier, occurrence of Japanese encephalitis (JE),
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