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. 2018 Sep 15;1095:112-118.
doi: 10.1016/j.jchromb.2018.07.017. Epub 2018 Jul 20.

Quantification of Hypoglycin A and Methylenecyclopropylglycine in Human Plasma by HPLC-MS/MS

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Quantification of Hypoglycin A and Methylenecyclopropylglycine in Human Plasma by HPLC-MS/MS

Aimee A Sanford et al. J Chromatogr B Analyt Technol Biomed Life Sci. .
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Hypoglycin A (HGA) and methylenecyclopropylglycine (MCPG) are naturally-occurring amino acids known to cause hypoglycemia and encephalopathy. Exposure to one or both toxins through the ingestion of common soapberry (Sapindaceae) fruits are documented in illness outbreaks throughout the world. Jamaican Vomiting Sickness (JVS) and seasonal pasture myopathy (SPM, horses) are linked to HGA exposure from unripe ackee fruit and box elder seeds, respectively. Acute toxic encephalopathy is linked to HGA and MCPG exposures from litchi fruit. HGA and MCPG are found in several fruits within the soapberry family and are known to cause severe hypoglycemia, seizures, and death. HGA has been directly quantified in horse blood in SPM cases and in human gastric juice in JVS cases. This work presents a new diagnostic assay capable of simultaneous quantification of HGA and MCPG in human plasma, and it can be used to detect patients with toxicity from soapberry fruits. The assay presented herein is the first quantitative method for MCPG in blood matrices.

Keywords: Acute toxic encephalopathy; Hypoglycin A (HGA); Methylenecyclopropylglycine (MCPG); Sapindaceae; Soapberry.


Figure 1
Figure 1. Varying SPE elution strength
Effects of varying the % acetonitrile in elution solvent on (A) recovery of HGA (gray squares) and MCPG (black circles), (B) extracted ion chromatograms of HGA with 30% (black solid) and 98% (gray dashed) acetonitrile elution, and (C) extracted ion chromatograms of MCPG with 30% (black solid) and 98% (gray dashed) acetonitrile elution.
Figure 2
Figure 2. Extracted ion chromatogram peak heights of spiked human plasma
Chromatograms of dns-HGA (left) at concentrations of (A) 0.00 ng/mL or unspiked, (B) 1.00 ng/mL, and (C) 150. ng/mL. Chromatograms of dns-MCPG (right) at concentrations of (D) 0.00 ng/mL or unspiked, (E) 5.00 ng/mL, and (F) 100. ng/mL. The MS/MS quantitation transitions shown dns-HGA and dns-MCPG are m/z 375.1→170.1 and m/z 361.1→157.1, respectively.
Figure 3
Figure 3. Evaluation of plasma matrix effects
(A) A 4.0 μL injection of processed pooled plasma in 0.1% formic acid in water was injected onto the C18 column. During injection, a solution of dns-HGA (top, gray, dashed) and dns-MCPG (bottom, black, solid) was infused post-column. No significant plasma matrix effects were observed at the expected retention times of 4.28 ± 0.03 minutes for dns-HGA and 3.88 ± 0.03 minutes for dns-MCPG. (B) For comparison, the same experiment was performed, with 4.0 μL of solvent blank (0.1% formic acid) injected onto the column during the infusion.
Scheme 1
Scheme 1
Derivatization of (A) HGA and (B) MCPG with dansyl chloride. Asterisks indicate 13C and 15N label sites on the internal standards.

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