Development of a novel method for analysing N-acetyl-DL-leucine enantiomers in human fingernail by UPLC-ESI-MS/MS and the evaluation in diabetes mellitus.

BACKGROUND: Recent research has been reported that N-acetyl-leucine content is significantly reduced in the saliva of diabetic patients, but no reports of detection in human nails have been found. This study aims to develop a novel method for the chiral separation of N-acetyl-DL-leucine (Ac-DL-Leu) in human fingernails to investigate the differences between healthy volunteers (HVs), prediabetes (PDs) and diabetic patients (DPs), and to verify its effectiveness in early warning of diabetes. METHOD: Chiral resolution was performed using DBD-Apy pre-column derivatization on a C18 column (2.1 × 150 mm, 1.9 µm) at 40 °C, and detected by UPLC-ESI-MS/MS. RESULTS: The resolution and the limit of detection (LOD) of Ac-DL-Leu were 1.75 and 1.50 fmol, respectively. The linear range of Ac-DL-Leu was 10-2000 fmol and the determination coefficient (R2) was above 0.9997. The recovery of Ac-DL-Leu in human nails was 96.92-105.69 %. The contents of Ac-D-Leu and Ac-L-Leu were analyzed in 18 HVs, 13 PDs and 16 DPs fingernails. The results showed that their contents were significantly lower in DPs than in PDs and HVs (p < 0.0001). CONCLUSIONS: A method for evaluating the effectiveness of Ac-DL-Leu enantiomers in human fingernails as a biomarker for diabetes was firstly developed.

The Effect of N-Acetyl-DL-Leucine on Neurological Symptoms in a Patient with Ataxia-Telangiectasia: a Case Study.

Ataxia-telangiectasia (AT) is a rare autosomal recessive disorder with no available curative treatment. Although the positive effects of N-acetyl-DL-leucine on cerebellar ataxia have been reported previously, there is little evidence of N-acetyl-DL-leucine's effects in patients with AT. This study assessed the effect of 16 weeks N-acetyl-DL-leucine supplementation on ataxia symptoms in a 9-year-old female with AT. The subject consumed 4 g/day N-acetyl-DL-leucine (2 g in the morning and 2 g in the evening) for 16 weeks. Safety was assessed via clinical blood chemistry prior to the intervention and after 6 and 16 weeks. Additionally, The Scale for the Assessment and Rating of Ataxia (SARA) score was used to assess the drug's effects on ataxia symptoms at baseline, 6, 12, and 16 weeks. Quality of life has also been evaluated by a specialist using the PedsQL questionnaire.Despite some initial (first week only) nausea and constipation, supplementation with N-acetyl-DL-leucine was well tolerated and safe according to blood chemistry measures. The SARA score progressively improved, and by week 16 had improved by 11.0 points (48.88%). Parent and self-reported quality of life assessments indicated physical, emotional, social, and school functions all improved by 16 weeks. Supplementation with N-acetyl-DL-leucine at a dose of 4 g/day for 16 weeks was well tolerated and significantly improved ataxia symptoms and quality of life measures in a young child with AT.

Determination of N-acetyl-DL-leucine in the saliva of healthy volunteers and diabetic patients using ultra-performance liquid chromatography with fluorescence detection.

BACKGROUND: Recent studies have indicated that N-acetyl-leucine (N-Ac-Leu) is a potential biomarker of diabetes. This study aimed to measure the levels of enantiomers of the chiral molecule N-Ac-DL-Leu in the saliva of patients with type 2 diabetes and further determine the potential association between them. METHOD: A novel validated method was established using ultra-performance liquid chromatography (UPLC) with fluorescence (FL) detection, in which precolumn derivatization of (R)-(-)-4-(N, N-dimethylaminosulfonyl)-7-(3-aminopyrrolidin-l-yl)-2,1,3-benzoxadiazole [(R)-(-)-DBD-APy] was used for the simultaneous determination and chiral separation of N-Ac-DL-Leu in human saliva. RESULTS: The labeled N-Ac-DL-Leu diastereomers were completely separated, with a resolution value of 1.93. Additionally, excellent linearity for N-Ac-DL-Leu was observed, with high coefficients of correlation (r2 ≥ 0.9999) in the range of 10-300 µM; the limit of quantitation (signal-to-noise ratio = 10) was 40-120 pmol/mL, and the mean recoveries of N-Ac-L-Leu and N-Ac-D-Leu were 102.48% and 104.68%, respectively. The levels of N-Ac-Leu in the saliva of diabetic patients and healthy volunteers were determined, and it was found that the levels of N-Ac-DL-Leu in the saliva of diabetic patients were significantly lower than those in healthy volunteers. (p < 0.01). CONCLUSIONS: The proposed method was successfully applied for the measurement of N-Ac-DL-Leu enantiomers in the saliva of diabetic patients and healthy volunteers.

Comparative analysis of pharmacological treatments with N-acetyl-DL-leucine (Tanganil) and its two isomers (N-acetyl-L-leucine and N-acetyl-D-leucine) on vestibular compensation: Behavioral investigation in the cat.

Head roll tilt, postural imbalance and spontaneous nystagmus are the main static vestibular deficits observed after an acute unilateral vestibular loss (UVL). In the UVL cat model, these deficits are fully compensated over 6 weeks as the result of central vestibular compensation. N-Acetyl-dl-leucine is a drug prescribed in clinical practice for the symptomatic treatment of acute UVL patients. The present study investigated the effects of N-acetyl-dl-leucine on the behavioral recovery after unilateral vestibular neurectomy (UVN) in the cat, and compared the effects of each of its two isomers N-acetyl-L-leucine and N-acetyl-D-leucine. Efficacy of these three drug treatments has been evaluated with respect to a placebo group (UVN+saline water) on the global sensorimotor activity (observation grids), the posture control (support surface measurement), the locomotor balance (maximum performance at the rotating beam test), and the spontaneous vestibular nystagmus (recorded in the light). Whatever the parameters tested, the behavioral recovery was strongly and significantly accelerated under pharmacological treatments with N-acetyl-dl-leucine and N-acetyl-L-leucine. In contrast, the N-acetyl-D-leucine isomer had no effect at all on the behavioral recovery, and animals of this group showed the same recovery profile as those receiving a placebo. It is concluded that the N-acetyl-L-leucine isomer is the active part of the racemate component since it induces a significant acceleration of the vestibular compensation process similar (and even better) to that observed under treatment with the racemate component only.