Lactulose Concentrations Research Articles

Quantitative analysis of lactose and lactulose in urine by high performance liquid chromatography for determination of intestinal lactase activity

Determination of intestinal lactase activity is directly done by measuring its activity in intestinal epithelium. This is an invasive method and ethically can not be done in healthy infants. Indirectly, determination of lactase activity, stated as excretion and ingestion ratio of lactose and lactulose, needs 30 hours hospitalized infants. The aim of this study was to look for a method for determination of lactase activity which is not invasive and not necessary hospitalized. Using this method lactose and lactulose were given as a single oral load after 2 hours fasting. Urine were collected for 5 hours starting from consuming sugar solution and then lactose and lactulose concentration in the urine were measured by High Performance Liquid Chromatography. The results showed single oral load of lactose and lactulose can be used for determination of lactase activity in infant and the infants were observed only for 7 hours. (Med J Indones 2003; 12: 8-12) Keywords: Lactase activity, Lactulose, Lactose, High Performance Liquid Chromatography

Available lysine, protein digestibility and lactulose in commercial infant formulas

Available lysine, in vitro protein digestibility and lactulose values were determined in 23 commercial infant formulas. The mean available lysine content of the formulas based on dairy proteins was 66.7±9.5 mg g−1 protein, similar to that of human milk, while that of soy based formulas was considerably lower (45.0±8.3 mg g−1 protein). In vitro protein digestibility values ranged 85.5–88.9% for soy-based formulas and 90.5–98.3% for formulas based on dairy proteins. Formulas based on milk enriched with whey had higher lactulose content than those based on cow's milk. However, all values were below the limit of 600 mg L−1 recommended for UHT milk.

Determination of lactulose and furosine in milk using front-face fluorescence spectroscopy

A novel rapid and non-destructive front-face fluorescence method to quantify furosine and lactulose in heat-treated milk has been developed. The emission fluorescence spectra (305-450 nm) of tryptophans of proteins, and emission and excitation fluorescence spectra of fluo- rescent Maillard-reaction products were recorded directly on milk samples. Principal component analysis (PCA) and principal component regression (PCR) were applied to process the spectra ob- tained. The front-face fluorescence method can give valuable information on the quality of heat- treated milk because it allows the evaluation of furosine and lactulose content simultaneously. The re- sults were well correlated with those of the reference methods. The front-face fluorescence technique is relatively low-cost, rapid and non-destructive and it could replace existing conventional analytical techniques practiced for lactulose and furosine evaluation in dairy products. Milk / heat treatment / front-face fluorescence / tryptophan / Maillard-reaction products / furosine / lactulose

Effects of early enteral feeding on the prevention of enterogenic infection in severely burned patients

The aim of the study was to analyse the effects of early enteral feeding on the prevention of enterogenic infection in severely burned patients. A total of 22 patients with severe burns were randomly divided into an early enteral feeding group (EF) and a delayed enteral feeding group (DF). The levels of serum endotoxin and TNF-α were dynamically detected in the members of both groups, and two unmetabolized sugars (lactulose and mannitol) were orally administered to these patients 1, 3 and 5 days postburn. Intestinal permeability was evaluated by detecting the concentrations of lactulose and mannitol in the urine and the lactulose-mannitol ratio (L/M) ratio. The levels of serum endotoxin and TNF-α in severely burned patients were significantly higher than in normal subjects ( P<0.01). The endotoxin level was positively related to the TNF-α level ( r EF=0.93, P<0.01; r DF=0.80, P<0.05). The urinary lactulose levels in both groups were significantly higher than in normal ( P<0.01), the urinary mannitol levels showed no obvious changes ( P>0.05). The urinary L/M ratios in both groups were significantly higher than in normal subjects ( P<0.01). The urinary L/M ratio was positively related to the serum endotoxin level ( r=0.95, P<0.01). The urinary lactulose levels and the urinary L/M ratios in the EF group were significantly lower than in the DF group ( P<0.01). The levels of serum endotoxin and TNF-α in the EF group were significantly lower than in the DF group ( P<0.01). It is suggested that intestinal permeability was markedly higher after burns than normal, and was positively related to the gut-derived endotoxemia. Early enteral feeding may decrease intestinal permeability, preserve the intestinal mucosal barrier and have a beneficial effect on the reduction of enterogenic infection.

Lactulose-mannitol intestinal permeability test: a useful screening test for adult coeliac disease.

The aim of this study was to determine the value of the lactulose mannitol intestinal permeability test in screening the general adult population for unrecognized enteropathy and latent coeliac disease. Subjects with positive serology (identified by screening carried out by the Belfast MONICA Project) along with controls were followed-up after 3 years and classified as having transient serology, persistent serology or coeliac disease. A 5-h urine collection was performed following the ingestion of 5 g lactulose, 2 g mannitol and glucose as an osmotic filler. Urinary concentrations of lactulose and mannitol were measured by enzymatic analysis. Percentage lactulose excretion (%LE) (0.94 versus 0.31, P<0.001) and lactulose mannitol excretion ratio (LMER) (0.12 versus 0.02, P<0.001) were significantly higher in screening-detected coeliac disease subjects compared with MONICA controls. The sensitivity of the permeability test was 87% in the screening situation compared with 81% in the clinical situation. In subjects with persistent and transient serology the LMER did not differ significantly from that of controls. The lactulose-mannitol test is a useful test for screening the general adult population for coeliac disease. Subjects with persistent and transient serology did not differ from MONICA controls and are unlikely to have latent coeliac disease.

Method development for the quantitative determination of lactulose in heat-treated milks by HPAEC with pulsed amperometric detection.

A robust, rapid, and sensitive high-performance anion-exchange chromatographic method for the separation and quantitative determination of lactulose in heated milks, along with other common milk carbohydrates, has been developed. Complete separation of galactose, glucose, N-acetylgalactosamine, lactose, lactulose, and epilactose was isocratically accomplished in about 22 min by an anion-exchange column eluted with 10 mM NaOH spiked with 2 mM Ba(OAc)2. The within-day repeatability was lower than 2.1% for 10 repetitive injections. Under optimized conditions, there was no need either of post-column addition of strong bases to the eluent for enhancing detection sensitivity or, even more important, for column regeneration between chromatographic runs. Upon 100-fold sample dilution, the amperometric response of lactulose in milk samples was found to be linear up to 100 microM (r = 0.99935) with a limit of detection equal to 1.2 microM (S/N = 3). The lactulose content in ultrahigh-temperature (UHT) and sterilized milks was evaluated by a calibration graph using 2-deoxyglucose as the internal standard, making the proposed method very useful in discriminating among heat-treated milks. Whereas the mean value of lactulose in skimmed, partially skimmed, and whole UHT milks ranged from 10 to 90 mg/100 mL, lactulose content in bottle-sterilized whole milk (two samples) was higher than 140 mg/100 mL. The presence of epilactose, which is another isomer of lactose, was also ascertained in sterilized milk.

Epidemiology of altered intestinal permeability to lactulose and mannitol in Guatemalan infants.

Subclinical alterations of small intestinal function have been reported frequently in tropical countries. Studies of small intestinal permeability to lactulose and mannitol were therefore completed in Guatemalan infants from a low-income, periurban community to assess the prevalence of altered intestinal function and the factors associated with this condition. Two hundred studies were successfully completed in 158 infants who had been free of diarrhea for at least 1 week before the day of study. Urinary concentrations of lactulose and mannitol during the 5-hour period after ingestion of 400 mg/kg body weight of lactulose and 100 mg/kg body weight of mannitol were measured by gas-liquid chromatography and compared by age group, feeding practices, anthropometric indexes, and serum iron and zinc concentrations. The overall prevalence of altered intestinal permeability (defined as a ratio of urinary recovery of lactulose to mannitol [L/M] > or =0.07) was 30%. The urinary L/M recovery ratio was positively associated with age; low weight for age; and, in infants less than 6 months of age, non-breast-feeding. Children with serum iron concentrations less than 7.16 microM/l (40 [microg/dl) had higher median L/M ratios (L/M = 0.068; 95% confidence interval [CI], 0.054, 0.085) than those with iron levels higher than this cutoff (L/M = 0.052; CI = 0.046, 0.058; p = 0.038). The median urinary L/M recovery ratio in 10 currently asymptomatic infants who had diarrhea during the week before testing (0.087; CI = 0.49, 0.154) was higher than that in children who had been free from diarrhea for at least 1 week (0.052; CI = 0.048, 0.056; p = 0.01). Age, feeding practices, low weight-for-age, low serum iron concentration, and recent diarrhea were all associated with altered intestinal function in this group of Guatemalan infants.

Screening procedure for evaluating heat load in commercial milks

The evaluation of heat damages in commercial milk samples has been studied by an ana lytical procedure which uses two different and complementary methods. The first is the rapid total sul- phydryl (SH) group measurement which evaluates whey protein denaturation. The second involves the measurement of absorbance at 340 nm (A340) of milks modified with the Clarifying Reagent', At .this wavelength sorne coloured compounds resulting from the non-enzymic browning still absorb. There is a satisfying correlation between A340 and lactulose content (r =0.98) measured in commercial milk samples by using capillary electrophoresis or high performance liquid chromatography (HPLC). Our analytical procedure using SH and A340 measurements has been applied to 130 raw and commercial milk samples. Analysis by Student's r-test shows that the values of the two parameters obtained for raw and heat-treated milk samples were significantly different (P < 0.01). This study demonstrates that this simple protocol makes possible rapid evaluation of heat damages in commercial milk sam- pies as il could be done with official but more sophisticated methods. © Inra/Elsevier, Paris. heat damage / commercial milk / sulphydryl group / absorbance A340 / transparent-modified miIk / analytical procedure

Low-Salt Water Reduces Intestinal Permeability in Atopic Patients

Background: Clinical remission of atopic dermatitis (AD) after intake of a low-salt water has recently been reported. Methods: We here investigated the effects of a low-salt water in AD patients with increased intestinal permeability to lactulose. From an initial group of 75 atopic patients, who undertook a treatment in a medical spa center, 12 were included because of abnormal urinary excretion of lactulose (>0.6%), which is absorbed in the enterocytic interspaces, at baseline. The excretion of mannitol with an extraenterocytic absorption was also studied as control. During 18 days, the patients followed a medical spa treatment course, including topical application and oral ingestion of a low-mineral water. Patients were asked not to modify their usual food regimen during the cure. At the end of treatment (day 18), the determination of the excretion of both sugars in urine was repeated. Results: At baseline, the mean urine excretion of lactulose in the study population was 1.21 ± 0.08 SEM. At the end of treatment (day 18), the lactulose concentration in urine diminished dramatically by 55% (mean 0.55 ± 0.09 SEM, p = 0.0005). Urine mannitol was also found to be reduced over the cure period, although to a lesser extent (30%, p = 0.04). Conclusion: This study indicates that low-mineral water normalizes the intestinal permeability of patients with AD.

Luminal Bacteria and Small-Intestinal Permeability

Background: The influence of luminal bacteria on small-intestinal permeability has not been fully assessed. This study addressed this issue. Methods: Thirty-four subjects (mean age 64 years; range 22-95 years) were investigated for possible small-intestinal bacterial overgrowth (SIBO) with culture of a small-intestinal aspirate. A lactulose/mannitol small-intestinal permeability test was performed, small-intestinal histology assessed and serum vitamin B12 concentrations measured in all subjects. Permeability was also assessed in a control group of 34 asymptomatic volunteers. Results: Urinary lactulose/mannitol ratios were significantly increased in subjects with SIBO with colonic-type flora (JP < 0.0005), even in the absence of villous atrophy. Urinary lactulose/mannitol ratios were increased in this group due to significantly increased urinary lactulose concentrations (P < 0.0005) rather than reduced urinary mannitol levels, after correcting for inter-subject variations in renal function. Counts of intraepithelial lymphocytes of CD8 phenotype were significantly increased in this group (P = 0.003). Although a significant correlation was found between intraepithelial lymphocyte counts and small-intestinal permeability overall (P < 0.002), these counts were not significantly different in subjects with SIBO with colonic-type flora whose permeability values were ≤ or ≥0.028, the upper limit of normal in asymptomatic controls. Serum vitamin B12 concentrations did not differ significantly between groups (P > 0.5). Ageing did not independently influence small-intestinal permeability (P>0.5). Conclusions: Small-intestinal permeability is increased in subjects with SIBO with colonic-type bacteria. This effect is independent of ageing and not mediated by vitamin B12 deficiency. Although counts of intraepithelial lymphocytes of CDS phenotype are increased in this disorder, it is also unlikely that these cells play an important causative role in this process. Routine light microscopic assessment underestimates the prevalence of small-intestinal functional disturbance in this disorder.

Correlation between Lactulose and Furosine in UHT-Heated Milk

The kinetics of furosine and lactulose formation during heat treatment of milk and correlations between these parameters have been studied as a contribution for setting legal standards for heat-processed milks. Activation energies of 93.15±5.87 kJ/mol and 153.2±6.49 kJ/mol for furosine and lactulose respectively were computed from zero-order rate constants. Polynomial and logarithmic equations were developed to predict the furosine content as a function of lactulose content. The polynomial equation proposed fit the experimental data and previously reported data. The standard error of the prediction was lower than ±7.8mg/liter. The application of this equation to observed values for lactulose and furosine in commercial milk samples would allow the detection of milk powder in UHT milk.

Lactulose, monosaccharides and undenatured serum protein contents in commercial UHT creams and their usefulness for thermal treatment assessment

Fifteen UHT cream samples with different fat levels (ranging from 10% to 35%) were analysed for free carbohydrates (lactose, lactulose, glucose, galactose and myoinositol) and undenaturated whey proteins using gas chromatography and high-performance liquid chromatography, respectively. Lactulose concentrations ranged from 7 to 177 mg per 100 ml, α-lactalbumin from 5.1 to 61.6 mg per 100 ml and β-lactoglobulin from 1.0 to 64.0 mg per 100 ml. These values were similar to those reported for UHT milks. The ratio β-lactoglobulin/lactulose was found to be significantly higher than that reported for UHT milk.

Automated determination of lactulose in milk using an enzyme reactor and flow analysis with integrated dialysis

A highly simplified and rapid enzymatic procedure for the determination of lactulose was developed. Lactulose was hydrolysed to fructose and galactose by β-galactosidase, and the amount of fructose was determined by using immobilized fructose dehydrogenase and K 3[Fe(CN) 6]as mediator. The reduced mediator was re-oxidised at a screen-printed Pt-electrode at a potential of 385 mV vs. a screen-printed Pt-pseudo reference electrode. An automated flow analysis (FA) method was developed allowing in-line hydrolysis of lactulose by soluble β-galactosidase from Aspergillus oryzae and separation of the analyte from the milk matrix by a dialysis unit. Optimised choice of reaction temperature (50°C) and reaction time (210 s) led to the applicability of the device to ultra-heat treated (UHT) and pasteurized milk samples without sample pretreatment. Good correlation of data to the official photometric test was obtained when the standard addition method was used to calculate lactulose concentrations. The automated FA system allowed a frequency of analysis of 17 samples h −1, compared to at least 12 h analysis time for the determination of the lactulose concentration in one milk sample with the German official method.

Measurement of sugar probes in serum: an alternative to urine measurement in intestinal permeability testing

The percentage dose of lactulose and mannitol excreted in urine after oral ingestion is used as a noninvasive method of assessing small intestinal permeability. The collection of incomplete or inaccurately timed urine samples can lead to errors in estimation of sugar probe molecules. We describe an HPLC method for the simultaneous determination of lactulose and mannitol in serum after oral ingestion of test sugars. We applied the test to healthy volunteers and to subjects undergoing jejunal biopsy for suspected gluten-sensitive enteropathy. The ratio of concentrations of lactulose and mannitol in serum discriminated well between subjects with a normal biopsy and those with villous atrophy, discrimination being best at 90 min postdose. The results agree well with lactulose:mannitol ratios determined in urine (r= 0.88), and the two methods can be used interchangeably. The determination of mannitol and lactulose in serum provides an acceptable alternative to urine collection and may be particularly useful in young children. It also reduces the time spent on the investigation from 5 h to 90 min.

Rapid and simultaneous quantification of rhamnose, mannitol, and lactulose in urine by HPLC for estimating intestinal permeability in pediatric practice

Determination of intestinal permeability by measuring nonmetabolized sugars has been used to assess the integrity of intestinal mucosa. We have developed and validated a modified HPLC method for determining the concentration of L-rhamnose, mannitol, and lactulose in urine, using an amine-modified silica column and refractive index detection. Probe sugars are simultaneously resolved within 18 min. The calibration curve for each sugar is linear to 20 mmol/L. The minimum detectable concentration of lactulose is 0.05 mmol/L. Recovery of probe sugars is between 99.3% and 105.1%. Overall precision (CV) of estimation of probe sugars ranges from 4.2% to 6.5%. In 14 urine samples from healthy children who ingested the test solution containing 1 g of L-rhamnose, 1 g of mannitol, and 5 g of lactulose, the 5-h urinary excretion ratios of lactulose/rhamnose and lactulose/mannitol were 0.047 +/- 0.018 and 0.021 +/- 0.010 (mean +/- SD), respectively. This method presents a rapid, convenient, and practical technique for determining intestinal permeability in clinical pediatric practice.

Evaluation of two methods for the determination of lactulose in milk

Two conventional methods for the determination of lactulose in milk were compared, namely the colorimetric-enzymatic method and the gas chromatographic method. Increasing lactulose concentrations were added to raw milk samples. For concentrations of lactulose higher than 50 mg L −1 both methods had a similar accuracy and precision. However, the enzymatic method was more sensitive and had a detection limit of about 10 mg L −1 . The detection limit for the gas chromatographic method was between 50 and 100 mg L −1 . In contrast to the gas chromatographic method, the enzymatic method can be used to discriminate between UHT-milk and high-pasteurized milk.

Evaluation of intestinal permeability in patients with inflammatory bowel disease using lactulose and measuring antibodies to lipid A.

This study looked at the intestinal permeability and the immune response to enteric bacterial antigens in patients with inflammatory bowel disease (IBD). They were evaluated by using a lactulose tolerance test and measuring blood anti-lipid A antibody concentrations, respectively. The lactulose tolerance tests were performed 22 times in 14 patients with Crohn's disease (CD), 19 times in 12 patients with ulcerative colitis (UC), and 12 times in 12 healthy controls. Blood lactulose concentrations were measured after oral administration every two hours for eight hours, also blood C reactive protein concentrations and anti-lipid A antibody concentrations were measured just before lactulose administration. Blood lactulose concentrations were significantly higher in patients with CD than in the controls from two to eight hours after administration, while in UC they were significantly higher than in the controls from six to eight hours. Maximum blood lactulose concentrations in each tolerance test in patients with the active phase significantly exceeded those in the inactive phase of either CD or UC. A significant correlation was also seen between the maximum blood lactulose concentrations and the C reactive protein concentrations. Blood anti-lipid A antibody concentrations in patients with CD were significantly higher than in the controls as well as in patients with UC in immunoglobulin (Ig) A class and IgG class. In UC they were significantly higher than in the controls in IgA class. But, they were not related to the severity of the disease of either CD or UC, and not correlated significantly with the maximum blood lactulose concentrations in either CD or UC. The intestinal permeability and the immune response to enteric bacterial antigens in patients with inactive CD were significantly increased over those in the controls as well as in patients with inactive UC. These findings suggest that an increase of the intestinal permeability and that of producing antibodies to enteric bacterial antigens are both important for the pathogenesis of IBD, and that the characteristics of CD and UC differ.

Effect of Absorption of D-Glucose and Water on Paracellular Transport in Rat Duodenum-Jejunum

Paracellular transport is thought to be a major absorptive pathway for small nutrient molecules. The authors used in vivo in situ perfusion of rat duodenum-proximal jejunum to examine paracellular transport using lactulose as a probe. They perfused solutions with a constant lactulose concentration but varied initial D-glucose concentration (range 12-176 mM) to open paracellular pathways and to increase water absorption, thereby optimizing potential for paracellular transport of lactulose and other solutes in its molecular weight range. All solutions contained sodium chloride to approach isotonicity. Water absorption was measured as the difference in weight of solution perfused and sample collected. Absorption of D-glucose increased with mean luminal D-glucose concentration, and water absorption more than doubled (from 0.12 +/- 0.03 to 0.26 +/- 0.05 mL/min per g dry wt of segment) as mean luminal glucose concentration was increased from 10 to 80 mM. Lactulose absorption was at the threshold of detection and did not correlate with D-glucose or water absorption. Expressed as percent per segment, D-glucose absorption ranged from 29-50%, and the lactulose absorption rate was 4-5%. The fraction of D-glucose absorption that could be attributed to lactulose absorptive pathways was 12% at the highest rate of water absorption. In conclusion, based on lactulose as a probe, under conditions of opening tight junctions by D-glucose, the paracellular component of D-glucose absorption was of the order of 1/10 of total D-glucose absorption (ie, not a major absorptive pathway.

Solubility of lactulose trihydrate

Lactulose (4-O-O-D-@aCtOpyranOSyl-D-fructose) was synthesized by Montgomery and Hudson [ 1] in 1930. Its physiological significance as a growth-promoting factor for Si$dobacterium (Bifidus Factor) was discovered by Petuely [ 21 in 1957. Bifidobacterium is a typically beneficial genus of the intestinal microflora and has useful functions for the host. Lactulose is now widely used as a Bifidus Factor in food, and as a remedy for the treatment of hepatic encephalopathy and chronic constipation in many countries of the world. Lactulose was crystallized first from alcohol solution as the anhydride [ 11. In 1992, Jeffrey et al. 131 examined lactulose trihydrate that had been crystallized from aqueous solution. The crystal structure and NMR analysis of lactulose trihydrate revealed that it was comprised only of /3-furanose, whereas lactulose anhydride showed the presence of /3furanose, a-furanose, and P-pyranose in the ratio of 75:10:X [ 41. Because lactulose trihydrate has been reported only recently, its physicochemical properties are not known in detail. The solubility of lactulose in water at 30, 60, and 90°C was reported by Oosten [ 51 and it showed a high solubility of above 76% w/w. However, information concerning the solubility profile of lactulose in aqueous solution at temperaturesbelow 30°C is not available. Since pharmaceutical products consisting of lactulose solutions with high lactulose concentrations are marketed worldwide, it is worthwhile to obtain further solubility information. This study examines the solubility of the crystal form of lactulose in aqueous solution at temperatures below 30°C.

Glutamine and the preservation of gut integrity

Parenteral glutamine dipeptide improves nitrogen balance in postoperative patients on total parenteral nutrition (TPM). Animal studies show that the structure and function of the gut is preserved by glutamine. It is not known if this is the case in human beings. 20 patients admitted to hospital for total parenteral nutrition were randomly allocated to receive parenteral nutrition enriched with glycyl-L-glutamine (Gln TPN), or standard parenteral nutrition (STPN). Mucosal biopsy specimens were taken from the second part of the duodenum before starting parenteral nutrition, and after two weeks. The ratio between the urine concentrations of lactulose and mannitol after enteral administration was used to measure intestinal permeability. After two weeks of parenteral nutrition in the GInTPN group, intestinal permeability was unchanged, whereas permeability in the STPN group increased. Villus height was unaltered in the GInTPN group but in the STPN group it decreased. The addition of glutamine to parenteral nutrition prevents deterioration of gut permeability and preserves mucosal structure.