Isotope-labeling derivatization with 3-nitrophenylhydrazine for LC/multiple-reaction monitoring-mass-spectrometry-based quantitation of carnitines in dried blood spots

Han, Jun; Higgins, Rehan; Lim, Mark D.; Atkinson, Kieran; Yang, Juncong; Lin, Karen; Borchers, Christoph H.

doi:10.1016/j.aca.2018.01.045

Cited by 30 publications

(16 citation statements)

References 27 publications

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“…For example, the strategies have been used to detect l -carnitine and its chiral isomers d -carnitine [60]. And just recently, an isotope-labelling strategy with 3-nitrophenylhydrazine as derivatization reagents was employed for LC-MS-based quantitation of acylcarnitines in dried blood spots with good linearity, high precision and high accuracy [85]. One important aspect to note is that the application of derivatization methods requires systematically methodological evaluation of the chemical stabilities of acylcarnitines under various reaction conditions before they are used in the practical sample analysis.…”

Section: Detection Of Acylcarnitines In Biological Samplesmentioning

confidence: 99%

Function, Detection and Alteration of Acylcarnitine Metabolism in Hepatocellular Carcinoma

2019

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Acylcarnitines play an essential role in regulating the balance of intracellular sugar and lipid metabolism. They serve as carriers to transport activated long-chain fatty acids into mitochondria for β-oxidation as a major source of energy for cell activities. The liver is the most important organ for endogenous carnitine synthesis and metabolism. Hepatocellular carcinoma (HCC), a primary malignancy of the live with poor prognosis, may strongly influence the level of acylcarnitines. In this paper, the function, detection and alteration of acylcarnitine metabolism in HCC were briefly reviewed. An overview was provided to introduce the metabolic roles of acylcarnitines involved in fatty acid β-oxidation. Then different analytical platforms and methodologies were also briefly summarised. The relationship between HCC and acylcarnitine metabolism was described. Many of the studies reported that short, medium and long-chain acylcarnitines were altered in HCC patients. These findings presented current evidence in support of acylcarnitines as new candidate biomarkers for studies on the pathogenesis and development of HCC. Finally we discussed the challenges and perspectives of exploiting acylcarnitine metabolism and its related metabolic pathways as a target for HCC diagnosis and prognosis.

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Section: Detection Of Acylcarnitines In Biological Samplesmentioning

confidence: 99%

Function, Detection and Alteration of Acylcarnitine Metabolism in Hepatocellular Carcinoma

2019

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“…This study applied 3NPH derivatization of acylcarnitines for comprehensive identification, as shown in a low-resolution quadrupole mass spectrometer. Derivatization by 3NPH [10] was used to increase the sensitivity and most importantly, to achieve a linear elution profile on a reversed phase column for all acylcarnitine classes, which is not the case for short acylcarnitines without derivatization [4].…”

Section: Resultsmentioning

confidence: 99%

“…An isotopically labeled internal acylcarnitine standard (IS, 20 pmol), containing 8 acylcarnitines species was added. Derivatization of acylcarnitines by 3-nitrophenylhydrazine (3NPH) was performed as previously reported [10]. In brief, 25 mM 3NPH (5 μL of 0.5 M in 35% acetonitrile), 25 mM EDC (2.5 μL of 1M in water), and 0.396% (0.4 μL, 99%) pyridine were added to the samples (in 80% methanol) sequentially and kept for 30 minutes at 30°C on a rocking platform.…”

Section: Methodsmentioning

confidence: 99%

“…Thus, all available acylcarnitine entries were extracted from the METLIN [8] database and the Human Metabolome Database (HMDB) [9], and a recent publication [6]. Acylcarnitines were derivatized by 3-nitrophenylhydrazine (3NPH), which modifies all carboxyl groups of acylcarnitines [10]. By adding a isotope-labeled 3NPH reference sample, a direct and unbiased comparison of two or more conditions is possible, since an identical metabolite extraction work-flow and LC-MS conditions are applied [11], but were not used in the current study.…”

Section: Introductionmentioning

confidence: 99%

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Acylcarnitine profiling by low-resolution LC-MS

Meierhofer

2019

PLoS ONE

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Acylcarnitines are fatty acyl esters of L-carnitine and facilitate the entry of long-chain fatty acids into mitochondria via the carnitine shuttle, where they are metabolized via ß-oxidation. Alterations of acylcarnitine species can be diagnostic for fatty acid oxidation disorders and organic aciduria and are thus frequently used to screen newborns. Only a subfraction of all known acylcarnitines is thereby monitored and quantified. Therefore, a method for the simultaneous fast and robust detection of all known acylcarnitines was developed using a single concise liquid chromatography mass spectrometry (LC-MS) approach. Derivatization by 3-nitrophenylhydrazine increased the signal intensity of the acylcarnitines and a linear elution from a reversed phase column was observed that was dependent on the length of the carbon chain. This allowed a precise prediction of the exact elution time for each acylcarnitine class, which depended solely on the chemical nature of the carbon chain. This method can be further used to screen for yet unknown acylcarnitine species and adds a layer of confidence for their correct identification. Altogether 123 acylcarnitines species were used to establish a targeted low-resolution LC-MS method. The method was applied to acylcarnitine profiling in several mouse tissues and fluids, in order to identify large differences in the quantity and composition of acylcarnitines.

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“…Carnitines were extracted from the dried blood spots in a two‐step liquid extraction procedure and then derivatized via a one‐pot reaction with 13 C 0 / 13 C 6 ‐NPH followed by UPLC–MS/MS in the MRM mode. With this method, 24 carnitines were monitored and limits of quantitation (LOQs) in the sub‐ to low‐femtomole levels were achieved (Han et al, ). Another hydrazine reagent used for the ICD of fatty acids is 2 H 0 / 2 H 20 ‐2,4‐bis(diethylamino)‐6‐hydrazino‐1,3,5‐triazine ( 2 H 0 / 2 H 20 ‐BDHT).…”

Section: Isotope‐coded Derivatization Reagents For Lc–ms Analysismentioning

confidence: 99%

Current trends in isotope‐coded derivatization liquid chromatographic‐mass spectrometric analyses with special emphasis on their biomedical application

El-Maghrabey

Kishikawa

Kuroda

2020

Biomedical Chromatography

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Currently, LC–MS has various applications in different areas such as metabolomics, pharmacokinetics, and pathological studies. Yet, matrix effects resulting from co‐existing constituents remain a major problem for LC–MS [or LC–tandem mass spectrometry (LC–MS/MS)]. Moreover, technical problems and instrumental drifts may lead to ion abundance variance. Thus, an internal standard (IS) is required to guarantee the accuracy and precision of the method. Because of their limited number, isotope‐coded derivatization (ICD) has been recently introduced to overcome this problem. For ICD, a stable heavy isotope‐coded moiety is used for labeling the standard or the control sample and the formed products can act as ISs. A light form of the reagent is used for labeling the sample. Then, both are mixed and analyzed by LC–MS(/MS). This strategy permits the identification of different unknown analytes including potential metabolites and disease biomarkers. All these attributes lead to persistent growth in the applications of ICD LC–MS(/MS) in various biomedical branches. In this article we review the ICD methods published in the last eight years for biomedical applications as well as briefly summarize other applications for environmental and food analyses as some of their used ICD reagents were further applied for analyzing biological specimens or have the potential for that.

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Isotope-labeling derivatization with 3-nitrophenylhydrazine for LC/multiple-reaction monitoring-mass-spectrometry-based quantitation of carnitines in dried blood spots

Cited by 30 publications

References 27 publications

Function, Detection and Alteration of Acylcarnitine Metabolism in Hepatocellular Carcinoma

Function, Detection and Alteration of Acylcarnitine Metabolism in Hepatocellular Carcinoma

Acylcarnitine profiling by low-resolution LC-MS

Current trends in isotope‐coded derivatization liquid chromatographic‐mass spectrometric analyses with special emphasis on their biomedical application

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