Simultaneous determination of dl-lactic acid and dl-3-hydroxybutyric acid enantiomers in saliva of diabetes mellitus patients by high-throughput LC–ESI-MS/MS

Abstract

A simultaneous determination method for the enantiomers of chiral carboxylic acids by the combination of ultraperformance liquid chromatography and mass spectrometry (UPLC-MS/MS) has been developed. (S)(+)-1-(2-Pyrrolidinylmethyl)-pyrrolidine (S-PMP) was used as the derivatization reagent for the high-throughput determination of biological chiral carboxylic acids, i.e., lactic acid (LA) and 3-hydroxybutyric acid (HA). The S-PMP efficiently reacted with the carboxylic acids under mild conditions at room temperature in the presence of 2,2'-dipyridyl disulfide and triphenylphosphine. The resulting S-PMP derivatives were highly responsive in the electrospray ionization (ESI)-MS operating in the positive-ion mode and gave characteristic product ions during the MS/MS, which enabled the sensitive detection using selected reaction monitoring. The derivatization was effective for the enantiomeric separation of the chiral carboxylic acids, and the resolution values of DL-LA and DL-HA were 4.91 and 9.37, respectively. Furthermore, a rapid separation of the derivatives of DL-LA and DL-HA within 7 min was performed using the UPLC system. The limits of detection on the column were in the low femtogram range (5-12 fg). The proposed procedure was successfully applied for the determination of the D- and L-isomers of LA and HA in the saliva of diabetes mellitus (DM) patients and healthy volunteers. The D-LA in DM patients was clearly higher than that in normal subjects. The derivatization followed by UPLC-ESI-MS/MS enabled the enantiomeric separation and detection of trace amounts of LA and HA in human saliva with a simple pretreatment and small sample volume.

Separation and determination of the enantiomers of lactic acid and 2‐hydroxyglutaric acid by chiral derivatization combined with gas chromatography and mass spectrometry

“…The results reveal that both the D-and L-LA levels in the plasma of T2DM mice were significantly (P < 0.05) higher than those of normal mice ( Figure 4A and B), which is consistent with the literature [30,31]. There are other studies that have reported that the L-LA levels increased slightly without significance [32,33]. Significant increases of D-LA were reported in all studies involving the LA enantiomers and T2DM [4][5][6][29][30][31][32], suggesting that D-LA may serve as an indicator for T2DM.…”

Establishment of a two-dimensional chiral HPLC system for the simultaneous detection of lactate and 3-hydroxybutyrate enantiomers in human clinical samples

“…7B), respectively. [27], and the %D-LA was 10.3%. The ratio was found to be 5.0% in our experiments.…”

“…However, it was presumably caused by the differences between individuals or by various bacteria in the saliva. However, our 2D-HPLC method for measuring D/L-3HB has better chromatographic separation efficiency than the LC-MS/MS method used by Tsutsui et al [27].…”

“…5). According to the results described above, a 2D-HPLC system combining a capillary monolithic ODS column and enantioselective columns (QD-AX and KSAACSP-001S) was established to separate the LA and 3HB enantiomers simultaneously.Toyo'oka et al report a LC-ESI-MS/MS method for the determination of LA and 3HB enantiomers recently[27]. A chiral amine, (S)(+)-1-(2-pyrrolidinylmethyl)-pyrrolidine (S-PMP) was used in their work as the derivatization reagent which refers to a so-called "indirect enantiomer analysis method.…”

Quantitation of α‐hydroxy acids in complex prebiotic mixtures via liquid chromatography/tandem mass spectrometry

“…However, previously reported analytical methods suffer from several innate shortcomings, including the use of time-consuming gas chromatography coupled to mass spectrometry (GC/MS), [17] the need for complex derivatization procedures with expensive reagents, [18] or, in some cases, using derivatization reagents that are potentially explosive. [19] Furthermore, more rapid approaches reported in the past have only demonstrated the detection of a limited number (1 to 5) of AHAs, [13,[20][21][22] or have not used more specific mass spectrometric detection. [23] Ideally, an improved AHA analytical method should be able to detect and quantitate a larger suite of AHAs in a single chromatographic run without derivatization.…”