Identification and quantification of the osmodiuretic mannitol in urine for sports drug testing using gas chromatography-mass spectrometry.

The osmodiuretic mannitol can be potentially misused in sports, owing to its urine diluting effect and the possibility to decrease bodyweight. To reveal a doping offence, resulting urinary mannitol concentrations after a prohibited intravenous application and a permitted oral intake have to be differentiated. Therefore, a reliable gas chromatography-mass spectrometry (GC-MS) method was established based on peracetyl derivatives of the analytes. All possible hexitols (allitol, galactitol, iditol, altritols, sorbitol and mannitol) that can occur in human urine were separated and identified on a phenyl-methylpolysiloxane column (HP-5MS) within 10.75 min, and the method demonstrated its capability for quantification purposes. The lower limit of detection and lower limit of quantification were estimated at 0.9 microg mL(-1) and 2.4 microg mL(-1), respectively, and the assay was validated for mannitol and sorbitol regarding the parameters specificity, linearity, intra- (<10%) and inter-day precision (<15%) and accuracy (92-102%). To investigate urinary mannitol concentrations after oral intake the method was applied to an excretion study, providing a mean urinary excretion of mannitol of 19.5%. Comparison of theoretically expected urinary levels after a common therapeutic dose of mannitol and preliminary results on physiological urinary mannitol levels were promising, regarding a threshold level for mannitol that can be utilised for doping control purposes.

Intestinal permeability is decreased in anorexia nervosa.

Malnutrition and absence of exogenous luminal nutrients in the gastrointestinal tract affect intestinal permeability (IP) leading to an increased penetration of substances that passively cross intestinal epithelium via intercellular pathways. We hypothesised that an increase in IP could occur in patients with anorexia nervosa because of their prolonged fasting and chronic malnutrition. Therefore, we assessed IP in 14 drug-free anorexic women and 19 drug-free age-matched healthy women by means of the lactulose/mannitol (LA/MA) test. To this purpose, after an overnight fast, subjects ingested an oral solution containing 5 g lactulose and 2 g mannitol in 100 ml water. Urine specimens were collected immediately before and 30, 60, 120, 180, 240 and 300 min after the ingestion of the sugar solution. Urinary lactulose and mannitol were determined by high-performance anion exchange chromatography coupled with pulsed amperometric detection. We found that IP, as expressed by the 5-h LA/MA excretion ratio, was significantly decreased in anorexic women because of a lower urinary recovery of lactulose. Moreover, in patients, the time course of lactulose excretion significantly differs from healthy controls. These results do not confirm our hypothesis of increased IP in anorexia nervosa. Since IP reflects the anatomo-functional status of intestinal mucosa, the present findings support the idea that changes in the anatomo-physiology of intestinal mucosa occur in anorexia nervosa.

Measurement of intestinal permeability using mannitol and lactulose in children with diarrheal diseases.

The excretion ratio of lactulose/mannitol in urine has been used to assess the extension of malabsorption and impairment of intestinal permeability. The recovery of lactulose and mannitol in urine was employed to evaluate intestinal permeability in children with and without diarrhea. Lactulose and mannitol probes were measured using high-performance liquid chromatography with pulsed amperometric detection (HPLC-PAD). Two groups of solutions containing 60 microM sugars were prepared. Group I consisted of glucosamine, mannitol, melibiose and lactulose, and group II of inositol, sorbitol, glucose and lactose. In the study of intra-experiment variation, a sample of 50 microl from each group was submitted to 4 successive determinations. The recovered amounts and retention times of each sugar showed a variation <2 and 1%, respectively. The estimated recovery was >97%. In the study of inter-experiment variation, we prepared 4 independent samples from groups I and II at the following concentrations: 1.0, 0.3, 0.1, 0.03 and 0.01 mM. The amounts of the sugars recovered varied by <10%, whereas the retention times showed an average variation <1%. The linear correlation coefficients were >99%. Retention (k'), selectivity (alpha) and efficiency (N) were used to assess the chromatographic conditions. All three parameters were in the normal range. Children with diarrhea presented a greater lactulose/mannitol ratio compared to children without diarrhea.

Measurement of intestinal permeability using mannitol and lactulose in children with diarrheal diseases

The excretion ratio of lactulose/mannitol in urine has been used to assess the extension of malabsorption and impairment of intestinal permeability. The recovery of lactulose and mannitol in urine was employed to evaluate intestinal permeability in children with and without diarrhea. Lactulose and mannitol probes were measured using high-performance liquid chromatography with pulsed amperometric detection (HPLC-PAD). Two groups of solutions containing 60 µM sugars were prepared. Group I consisted of glucosamine, mannitol, melibiose and lactulose, and group II of inositol, sorbitol, glucose and lactose. In the study of intra-experiment variation, a sample of 50 µl from each group was submitted to 4 successive determinations. The recovered amounts and retention times of each sugar showed a variation <2 and 1 per cent, respectively. The estimated recovery was >97 per cent. In the study of inter-experiment variation, we prepared 4 independent samples from groups I and II at the following concentrations: 1.0, 0.3, 0.1, 0.03 and 0.01 mM. The amounts of the sugars recovered varied by <10 per cent, whereas the retention times showed an average variation <1 per cent. The linear correlation coefficients were >99 per cent. Retention (k'), selectivity (a) and efficiency (N) were used to assess the chromatographic conditions. All three parameters were in the normal range. Children with diarrhea presented a greater lactulose/mannitol ratio compared to children without diarrhea.

Gut permeability in IgA nephropathy.

Gut permeability to small molecules was assessed by the differential absorption of cellobiose and mannitolin 18 patients with IgA nephropathy (IgAN). The urinary cellobiose:mannitol excretion ratio in patients did not differ (P = 0.42) from controls. These findings do not support the hypothesis that chronic increased mucosal permeability allows excessive antigen penetration to the mucosal immune system, predisposing to glomerular IgA deposition in IgA nephropathy. However, the patient with the greatest cellobiose:mannitol ratio developed macroscopic haematuria within 3 weeks of testing, raising the possibility of a transient abnormality in gut permeability.