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1.
J Mass Spectrom ; 54(11): 878-884, 2019 Nov.
Article in English | MEDLINE | ID: mdl-31652368

ABSTRACT

Overcoming the detrimental effects of sweet spots during crystallization is an important step to improve the quantitative abilities of matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. In this study, we introduce MALDI targets, which exhibit a channel design to reduce sweet spot phenomena and improve reproducibility. The size of the channels was 3.0 mm in length, 0.35 mm in depth, and 0.40 mm in width, adjusted to the width of the implemented laser beam. For sample deposition, the matrix/sample mixture was homogenously deposited into the channels using capillary action. To demonstrate the proof-of-principle, the novel plates were used for the quantification of acetyl-L-carnitine in human blood plasma using a combined standard addition and isotope dilution method. The results showed that the reproducibility of acetyl-L-carnitine detection was highly improved over a conventional MALDI-MS assay, with RSD values of less than 5.9% in comparison with 15.6% using the regular MALDI method. The limits of quantification using the new plates were lowered approximately two-fold in comparison with a standard rastering approach on a smooth stainless-steel plate. Matrix effects were also assessed and shown to be negligible. The new assay was subsequently applied to the quantification of acetyl-L-carnitine in human plasma samples.


Subject(s)
Acetylcarnitine/blood , Crystallization , Humans , Limit of Detection , Reproducibility of Results , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Stainless Steel/chemistry , Surface Properties
2.
Cell Metab ; 22(3): 472-84, 2015 Sep 01.
Article in English | MEDLINE | ID: mdl-26256392

ABSTRACT

Mitochondrial reactive oxygen species (ROS) play a central role in most aging-related diseases. ROS are produced at the respiratory chain that demands NADH for electron transport and are eliminated by enzymes that require NADPH. The nicotinamide nucleotide transhydrogenase (Nnt) is considered a key antioxidative enzyme based on its ability to regenerate NADPH from NADH. Here, we show that pathological metabolic demand reverses the direction of the Nnt, consuming NADPH to support NADH and ATP production, but at the cost of NADPH-linked antioxidative capacity. In heart, reverse-mode Nnt is the dominant source for ROS during pressure overload. Due to a mutation of the Nnt gene, the inbred mouse strain C57BL/6J is protected from oxidative stress, heart failure, and death, making its use in cardiovascular research problematic. Targeting Nnt-mediated ROS with the tetrapeptide SS-31 rescued mortality in pressure overload-induced heart failure and could therefore have therapeutic potential in patients with this syndrome.


Subject(s)
Heart Failure/metabolism , Mitochondria, Heart/metabolism , NADP Transhydrogenases/metabolism , NADP/metabolism , Oxidative Stress , Adenosine Triphosphate/metabolism , Animals , Cells, Cultured , Glutathione/metabolism , Heart Failure/pathology , Mice, Inbred C57BL , Mitochondria, Heart/pathology , Reactive Oxygen Species/metabolism
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