Time-based investigation of urinary metabolic markers for Type 2 diabetes: Metabolomics approach for diabetes management.

Diabetes is considered one of the most important health emergencies worldwide and Egypt has 8.2 million diabetic patients according to the International Diabetes Federation report in 2017. The objective of this study was to monitor the time-course variation in the metabolic profile of diabetic rats to detect urinary metabolic biomarkers using the metabolomics approach. Type 2 diabetes was induced in male Wistar albino rats using a single intraperitoneal injection of 40 mg/kg of streptozotocin following oral administration of 10% fructose in drinking water for 3 weeks. Then, urine was collected for 24 h from rats at three time points (0, 2, and 4 weeks after confirmation of diabetes), and were analyzed by nuclear magnetic resonance (H1 -NMR), followed by multivariate data analysis. The results from H1 -NMR pointed out that d-glucose, taurine, l-carnitine, l-fucose, 1,5-anhydrosorbitol, and d-galactose levels showed consistent significant variation (p < 0.05) between the positive (diabetic) and negative (normal) controls during the whole experimental period. Also, with the disease progression, myoinositol, and l-phenylalanine levels were significantly altered (p < 0.05) after 2 weeks and this alteration was maintained till the end of the 4-week experimental period in the positive control group. From the results of the present study, it could be concluded that we cannot depend only on glucose levels for prognostic purposes since there are other metabolic disturbances in diabetes which need to be tracked for better disease prognosis.

An in-depth Comparison of the Pediatric and Adult Urinary N-glycomes.

We performed an in-depth characterization and comparison of the pediatric and adult urinary glycomes using a nanoLC-MS/MS based glycomics method, which included normal healthy pediatric (1-10 years, n = 21) and adult (21-50 years, n = 22) individuals. A total of 116 N-glycan compositions were identified, and 46 of them could be reproducibly quantified. We performed quantitative comparisons of the 46 glycan compositions between different age and sex groups. The results showed significant quantitative changes between the pediatric and adult cohorts. The pediatric urinary N-glycome was found to contain a higher level of high-mannose (HM), asialylated/afucosylated glycans (excluding HM), neutral fucosylated and agalactosylated glycans, and a lower level of trisialylated glycans compared with the adult. We further analyzed gender-associated glycan changes in the pediatric and adult group, respectively. In the pediatric group, there was almost no difference of glycan levels between males and females. In adult, the majority of glycans were more abundant in males than females, except the high-mannose and tetrasialylated glycans. These findings highlight the importance to consider age-matching and adult sex-matching for urinary glycan studies. The identified normal pediatric and adult urinary glycomes can serve as a baseline reference for comparisons to other disease states affected by glycosylation.

Development of an amperometric biosensing method for the determination of L-fucose in pretreated urine.

The first amperometric biosensing method for the determination of L-fucose is described. L-Fucose is the objective of much current research, as it is considered as a potential marker for various pathologic disorders. Recombinant L-fucose dehydrogenase, having as cofactor beta-nicotinamide adenine dinucleotide phosphate (NAD+P), was cross-linked in a water-soluble photosensitive polymer matrix, that is, polyvinyl alcohol (PVA) modified with styrylpyridinium (SbQ), in the presence of BSA and glutaraldehyde. The resulting membrane was sandwiched between two polycarbonate membranes and was mounted in an amperometric cell. The oxidation of the enzymatically produced NADPH was monitored at a platinum anode at +0.25 V versus a silver pseudoreference electrode in the presence of ferricyanide. The system was fully optimized with respect to various analytical parameters. Regarding to the mechanical properties of the membrane and the storage stability of the immobilized enzyme, various parameters were also optimized. Several methods for the pretreatment of urine samples were investigated. Treatment of the samples with PbO2 found to eliminate the interference effect of various electroactive species exist in urine; optimum incubation time was determined since at prolonged incubation times L-fucose is also affected. Calibration curves for the direct and the mediated monitoring of NADPH were liner over the concentration ranges 0.04-1.0 mM (r2=0.9995) and 0.03-3.0 mM (r2=0.9997) fucose, respectively. The detection limits (S/N 3) were 2 and 1.5 microM fucose, respectively. The R.S.D. of the mediated biosensor is better than 1.5% (n=10, 0.5 mM fucose). The proposed biosensor correlates well with a reference enzymatic method and exhibits very good working and storage stability.

Combination assay of urinary free L-fucose and trypsin inhibitor may be useful indicators of disease activity in patients with hematologic malignancies.

L-Fucose is a monosaccharide located at the non-reducing ends of sugar chains of glycoconjugates. Urinary L-fucose (U-FC) is excreted as free L-fucose, and clinically useful as a tumor marker of digestive organ cancers. We evaluated the clinical usefulness of U-FC levels in patients with various hematologic malignancies because U-FC for hematologic malignancies has only rarely been described. The mean U-FC levels in the acute nonlymphocytic leukemia (ANLL), myelodysplastic syndromes (MDS) and myeloproliferative disorders (MPD) groups were significantly higher than in the control group (P < 0.05). Recently, we reported that urinary trypsin inhibitor (UTI) levels in patients with ANLL, MDS, non-Hodgkin's lymphoma and multiple myeloma were significantly elevated, compared with those in healthy adult volunteers. Noninvasive combination assays of UTI and U-FC may have a higher accuracy in diagnosis of ANLL and MDS than those of UTI or U-FC alone. UTI and U-FC combination assays, noninvasive for patients, could be expanded as useful indicators in hematologic malignancies.

[A study on the spectra of L-fucose].

The concentration of L-fucose will extraordinarily increase in urine in patients with early stage of cancer. It seems that this is a significant clue for early phase diagnosis of cancer. Unfortunately the sensitive detection of L-fucose is difficult. Thus we carried out researches on IR-spectrum, Raman spectrum, ultraviolet absorption spectrum and ultraviolet fluorescence spectrum. We compared the results from patients and healthy subjects. It is confirmed that the spectroscopic method, especially ultraviolet fluorescence spectrophotometry, may be suitable for early stage screening of some kinds of cancer. This test is rapid, inexpensive and painless.

Starch and dietary fiber components are excreted and degraded to variable extents in ileostomy subjects consuming mixed diets with wheat- or oat-bran bread.

The study was conducted to determine if the excretion of starch and dietary fiber components varies in ileostomy subjects consuming diets high or low in dietary fiber. Excretion of starch, enzyme-resistant starch and dietary fiber components was studied in nine human subjects with ileostomies, who consumed (in a crossover design) a wheat bread-based diet (daily intake 274 g starch, 2.4 g enzyme-resistant starch and 14.4 g total dietary fiber) and a high fiber diet based on oat-bran bread (daily intake 243 g starch, 2.7 g enzyme-resistant starch and 40.2 g total dietary fiber). Food and excreta were collected on d 3 and 17. No significant differences in excretion of starch, enzyme-resistant starch or dietary fiber components were found on these 2 d in each dietary period. When subjects consumed the wheat bread-based diet they excreted (mean +/- SD) 3.3 +/- 1.7 g starch and 2.4 +/- 0.4 g enzyme-resistant starch daily, whereas when consuming the oat bran-based diet they excreted 4.5 +/- 3.1 g starch and 2.5 +/- 0.4 g enzyme-resistant starch. During both dietary periods subjects excreted significantly greater amounts of certain dietary fiber polysaccharide residues (fucose, galactose and uronic acid) than they ingested. This indicates a contribution of endogenous and/or microbial material to the dietary fiber value in ileostomy effluents. However, significantly less excretion of some dietary fiber polysaccharide residues, especially glucose residues, during the oat-bran bread-based dietary period was also noted. This was presumably caused by a degradation of mixed-linked (1,3),(1,4)-beta-D-glucans.

Fucosidosis: una enfermedad por almacenamiento lisosomal / Fucosidosis: a lysosomal storage disease

Se describen 3 pacientes: 2 del sexo masculino y 1 del femenino, con enfermedad por almacenamiento lisosomal del tipo fucosidosis. En los 3 casos existió el antecedente de retardo psicomotor y posteriormente se hizo evidente el deterioro progresivo de las capacidades psíquicas motoras adquiridas. Todos tenían facies toscas, retardo mental profundo y alteraciones morfológicas en vértebras lumbares. Desde el punto de vista bioquímico, los 3 pacientes presentaron excreción urinaria de oligisacáridos ricos en fucosa, así como disminución de la actividad de la enzima alfa-L-fucosidasa en leucocitos. Se hace énfasis en la demostración del defecto enzimático como único criterio diagnóstico confirmatorio

Fucosidosis: una enfermedad por almacenamiento lisosomal / Fucosidosis: a lysosomal storage disease

Se describen 3 pacientes: 2 del sexo masculino y 1 del femenino, con enfermedad por almacenamiento lisosomal del tipo fucosidosis. En los 3 casos existió el antecedente de retardo psicomotor y posteriormente se hizo evidente el deterioro progresivo de las capacidades psíquicas motoras adquiridas. Todos tenían facies toscas, retardo mental profundo y alteraciones morfológicas en vértebras lumbares. Desde el punto de vista bioquímico, los 3 pacientes presentaron excreción urinaria de oligisacáridos ricos en fucosa, así como disminución de la actividad de la enzima alfa-L-fucosidasa en leucocitos. Se hace énfasis en la demostración del defecto enzimático como único criterio diagnóstico confirmatorio

Urinary level of L-fucose as a marker of alcoholic liver disease.

The urinary levels of L-fucose were measured in 93 alcoholics; 20 of these were without liver disease, 57 with noncirrhotic alcoholic liver disease, and 16 with alcoholic liver cirrhosis. In addition, patients with cirrhosis due to viral infection, and healthy subjects were evaluated. The mean urinary L-fucose concentration showed significantly higher values in patients with alcoholic liver disease and alcoholic liver cirrhosis when compared with the healthy subjects or the chronic alcoholics without liver disease (p < 0.001). The urinary L-fucose level was also significantly higher (p < 0.001) in cases of alcoholic liver cirrhosis than in noncirrhotic alcoholic liver disease (384 +/- 97 vs. 240 +/- 95 mumol/g of creatinine). No difference was observed between the healthy subjects and chronic alcoholics without liver disease (143 +/- 29 vs. 155 +/- 60 mumol/g of creatinine). The urinary level of L-fucose was significantly higher with alcoholic cirrhosis (384 +/- 97 mumol/g of creatinine) than with viral cirrhosis (265 +/- 42 mumol/g of creatinine) (p < 0.001). The measurement of urinary L-fucose may be a useful marker of alcoholic liver disease.

[Level of certain carbohydrate components in blood serum and urine of patients stomach and duodenal ulcer patients in the late post-vagotomy period]. / Soderzhanie nekotorykh uglevodnykh komponentov v syvorotke krovi i moche bol'nykh v otdalennye sroki posle vagotomii pri iazvennoi bolezni zheludka i dvenadtsatiperstnoi kishki.

Estimation of protein-bound hexoses, fucose and sialic acids in the blood and urine of 64 patients with gastroduodenal ulcers within later periods after various types of gastric vagotomy showed that the denervation inhibited the protective barrier of gastric and duodenal mucosal membrane. Protective functions of gastroduodenal mucosal membrane were most distinctly inhibited after selective proximal vagotomy which may be responsible for higher amounts of recurrences of ulcerous disease after this type of vagotomy than other types of denervation.

[Clinical assessment of urinary free L-fucose levels].

We measured urinary levels of free L-fucose in healthy subjects, patients with benign diseases, and patients with cancer using an automated analyzer and a newly isolated L-fucose dehydrogenase, and evaluated the clinical usefulness of the results. The values obtained were corrected for urinary creatinine as micromoles per gram of creatinine. The cutoff value, set at the mean + 2SD for the healthy subjects, was 250 mumol/g.Cr. Patients with gallbladder cancer, bile-duct cancer, liver cancer, pancreatic cancer, or cirrhosis of the liver had significantly higher levels of L-fucose than the healthy subjects. The diagnostic sensitivity for these five diseases, taken together, was 68% (144/213). Specificity for the detection of cancer was calculated by use of false positives for patients with cholelithiasis, hepatitis, and pancreatitis: it was 73% (76/104). Diagnostic accuracy for these seven diseases taken together was therefore 69% (220/317). We compared the positive ratio of the L-fucose level with that of the tumor markers AFD and CA19-9. The positive ratio of an L-fucose value above the cutoff was higher than the positive ratio of either marker in bile-duct cancer, gallbladder cancer, liver cancer, and pancreatic cancer. The results suggested that the urinary levels of free L-fucose reflected the metabolism of sugar chains of glycoconjugates, and may be usefully clinically as a tumor marker.

Assay of urinary free fucose by fluorescence labeling and high-performance liquid chromatography.

The concentrations of free fucose and other sugars in urine of cancer patients and healthy subjects were analyzed by high-performance liquid chromatography. After the urine samples were dried, we coupled the sugars in the residue with 2-aminopyridine to be detected as fluorescent derivatives. We then analyzed the pyridylamino derivatives of the sugars with an anion-exchange column and borate buffer. The difference between cancer patients and healthy subjects for the mean concentrations of fucose corrected for creatinine was significant (P less than 0.001). We checked the relationship between the concentrations of other sugars and the presence of cancer. This method is highly sensitive, and neither a cleanup procedure before labeling nor purification before injection into the column is needed. Not only fucose but also other sugars can be detected simultaneously, so this method should be useful for studying any changes in sugars in urine in various diseases.

Characterization and 400-MHz 1H-NMR analysis of urinary fucosyl glycoasparagines in fucosidosis.

Fucosyl glycoasparagines accumulating in the urine of a patient with fucosidosis were isolated using reverse-phase HPLC. Structural analysis of 25 glycoasparagines was carried out by combination of methylation and 400-MHz 1H-NMR spectroscopy analyses. The compounds represent different steps in the incomplete catabolism of N-glycosidically linked glycans, as the result of an alpha-L-fucosidase deficiency. All of the glycoasparagines possess a fucose residue alpha-1,6-linked to the GlcNAc 1 residue attached to asparagine. Fucose residues on the peripheral branches were linked either alpha-1,3 to GlcNAc residues (X determinant) or alpha-1,2 to galactose residues (H determinant). The present study allows precise assignments of the NMR parameters for most of the fucosyl linkages occurring in N-glycosidically linked glycans of the N-acetyllactosamine type.

Rapid, simple enzymatic assay of free L-fucose in serum and urine, and its use as a marker for cancer, cirrhosis, and gastric ulcers.

We devised a kit for use with automated analyzers, for assay of urinary free L-fucose by means of a newly isolated L-fucose dehydrogenase (EC 1.1.1.122), and we measured L-fucose in healthy subjects, cancer patients, and patients with other diseases. It takes 10 min to complete one assay. Absorbance and L-fucose concentration were linearly related up to at least 3.0 mmol/L, analytical recovery was 90-104%, and intra- and interassay coefficients of variation were less than 4.2% and 7.8%, respectively. The concentrations of L-fucose, corrected for creatinine, were significantly higher than those in healthy subjects in nine of 18 patients with gastric ulcers, 19 of 21 patients with cirrhosis of the liver, and 206 of 366 patients with some type of cancer, reflecting a changed L-fucose metabolism. Because urine specimens are analyzed and the test is rapid and inexpensive, this method may be suitable for mass screening for some kinds of cancer, cirrhosis, and gastric ulcers.