Plasma esterified and non-esterified fatty acids metabolic profiling using gas chromatography-mass spectrometry and its application in the study of diabetic mellitus and diabetic nephropathy.

Using gas chromatography-mass spectrometry (GC-MS), a new metabolic profiling method was established to assess the levels of non-esterified fatty acids (NEFAs) and esterified fatty acids (EFAs) in plasma. The extraction method was simple and robust without removing protein process. With this method 25 fatty acids (FAs), both EFAs and NEFAs, can be recognized simultaneously with only 10 µL plasma. 15 of the 25 can be precisely quantified. The method was validated and then applied into clinical metabonomics research. Five clinical groups including 150 cases were involved. The relationship between FA levels and diabetic mellitus (DM) as well as diabetic nephropathy (DN) pathology was speculated. Furthermore, the possible pathological causes and effects were discussed in detail. Potential biomarkers (p value <0.01) were screened with Student's t-test. With the application of partial least squares-discriminant analysis (PLS-DA), different stages were distinguished. The result may be useful for the pathology study of metabolic syndromes, and may also be helpful for monitoring the progression of DM and DN.

Correlations of six related pyrimidine metabolites and diabetic retinopathy in Chinese type 2 diabetic patients.

BACKGROUND: Diabetic retinopathy (DR), a kind of diabetic microvascular complication, is the leading cause of visual impairment in adults aged 30 to 65 years. Despite rapid research progress, robust predictors to assess prospectively with high precision the risk for DR in individuals with diabetes are still lacking. We investigated the relationship between pyrimidine metabolites and disease, and find out the potential biomarkers for diagnosis. METHODS: The study group consisted of 116 subjects who were divided to 3 groups: control (n=41), type 2 diabetes without retinopathy (DM, n=37), and with retinopathy (DR, n=38). Biochemical and clinical parameters, concentrations of related metabolites, including of cytosine, cytidine, uridine, thymine, thymidine and 2'-deoxyuridine were measured in plasma of all participants. RESULTS: There was a significant increase of concentrations of cytosine (p=0.010), cytidine (p<0.001) and thynidine (p<0.001) with DR compared to DM. The concentration of uridine, thymine and 2'-deoxyuridine did not change. CONCLUSIONS: The concentrations of cytosine, cytidine and thynidine may be useful for monitoring the progression of DR and evaluating the treatment. And cytidine has good sensitivity and specificity for diagnosis.

Correlations of creatine and six related pyrimidine metabolites and diabetic nephropathy in Chinese type 2 diabetic patients.

OBJECTIVES: The assessment of the clinical significance of creatine, cytosine, cytidine, uridine, thymine, thymidine, and 2'-deoxyuridine concentrations in patients with type 2 diabetes mellitus (T2DM) and diabetic nephropathy (DN) for the detection of the relationship between pyrimidine metabolites and disease. DESIGN AND METHODS: The study group consisted of 119 subjects, which were divided to three groups: control (n=31), type 2 diabetes without nephropathy (DM, n=23), and with nephropathy (DN, n=65). Levels of related metabolites were measured in plasma of all participants. RESULTS: There is a significant increase in levels of cytosine (P<0.001), cytidine (P<0.001), and thymidine (P=0.016) with DN compared to DM. The levels of uridine, thymine, 2'-deoxyuridine, and creatine did not change. CONCLUSIONS: The levels of cytosine, cytidine, and thymidine may be useful for monitoring the progression of DM and evaluating the treatment.

Ultraviolet and tandem mass spectrometry for simultaneous quantification of 21 pivotal metabolites in plasma from patients with diabetic nephropathy.

A sensitive and specific method was developed for simultaneous determination of 21 compounds related to the diabetic nephropathy (DN) in a single analysis using high-performance liquid chromatography coupled to ultraviolet and tandem mass spectrometry (HPLC-UV/MS/MS) in human plasma. With retention times and MS/MS for peak identification, both UV and MS detectors were used for quantification. Calibration curves suitable for the analysis of plasma were linear (r(2)>0.998) with limits of detection (LOD) from 10 to 1000 ng/mL. Intraday relative standard deviation (R.S.D.) and interday R.S.D. were both lower than 15%. With the case and control study, we found five potential biomarkers of DN, including adenosine, inosine, uric acid, xanthine and creatinine.

Simultaneous determination of sixteen metabolites related to neural tube defects in maternal serum by liquid chromatography coupling with electrospray tandem mass spectrometry.

Disturbances in maternal folate, homocysteine, and glutathione metabolism have been reported to be associated with neural tube defects (NTDs). However, the role played by specific components in the metabolic pathways leading to NTDs remains unclear. Thus an analytical method for simultaneous measurement of sixteen compounds involved in such three metabolic pathways by high performance liquid chromatography-tandem mass spectrometry was developed. The use of hydrophilic chromatography column improved the separation of polar analytes and the detection mode of multiple-reaction monitoring (MRM) enhanced the specificity and sensitivity so as to achieve simultaneous determination of three class of metabolites which have much variance in polarity and contents. The influence of parameters such as temperature, pH, flow rate on the performance of the analytes were studied to get an optimal condition. The method was validated for its linearity, accuracy, and precision, and also used for the analysis of serum samples of NTDs-affected pregnancies and normal women. The result showed that the present method is sensitive and reliable for simultaneous determination of as many as sixteen interesting metabolites which may provide a new means to study the underlying mechanism of NTDs as well as to discover new potential biomarkers.

Correlations of six related purine metabolites and diabetic nephropathy in Chinese type 2 diabetic patients.

OBJECTIVES: The assessment of the clinical significance of adenosine, adenine, inosine, xanthine, hypoxanthine and uric acid concentrations in patients with type 2 diabetes mellitus (T2DM) and diabetic nephropathy (DN) for the detection of the relationship between purine metabolites and disease. DESIGN AND METHODS: The study group consisted of 119 subjects which were divided into three groups: control (n=31), type 2 diabetes without nephropathy (DM, n=23) and with nephropathy (DN, n=65). Levels of related metabolites were measured in plasma of all participants. RESULTS: There is a significant increase of levels of adenosine (P<0.001), inosine (P<0.001), xanthine (P=0.012) and uric acid (P=0.016) with DN compared to DM. The level of xanthine oxidase (reflected by the uric acid: xanthine) did not change. CONCLUSION: The levels of adenosine, inosine, uric acid and xanthine may be useful for monitoring the progression of DM and evaluating the treatment.