Inulin increases the beneficial effects of rhubarb supplementation on high-fat high-sugar diet-induced metabolic disorders in mice: impact on energy expenditure, brown adipose tissue activity, and microbiota.

Consumption of prebiotics and plant-based compounds have many beneficial health effects through modulation of gut microbiota composition and are considered as promising nutritional strategy for the treatment of metabolic diseases. In the present study, we assessed the separated and combined effects of inulin and rhubarb on diet-induced metabolic disease in mice. We showed that supplementation with both inulin and rhubarb abolished the total body and fat mass gain upon high-fat and high-sucrose diet (HFHS) as well as several obesity-associated metabolic disorders. These effects were associated with increased energy expenditure, lower whitening of the brown adipose tissue, higher mitochondria activity and increased expression of lipolytic markers in white adipose tissue. Despite modifications of intestinal gut microbiota and bile acid compositions by inulin or rhubarb alone, combination of both inulin and rhubarb had minor additional impact on these parameters. However, the combination of inulin and rhubarb increased the expression of several antimicrobial peptides and higher goblet cell numbers, thereby suggesting a reinforcement of the gut barrier. Together, these results suggest that the combination of inulin and rhubarb in mice potentiates beneficial effects of separated rhubarb and inulin on HFHS-related metabolic disease and could be considered as nutritional strategy for the prevention and treatment of obesity and related pathologies.

Fusion and secretory expression of an exo-inulinase and a d-allulose 3-epimerase to produce d-allulose syrup from inulin.

BACKGROUND: This study developed a feasible catalytic method for d-allulose syrup production using a fusion enzyme, either in free or immobilized form, through hydrolysis of inulin extracted from Jerusalem artichoke tubers. RESULTS: d-Allulose 3-epimerase (DAE) was actively expressed in secretory form by fusing with the extracellular exo-inulinase CSCA in Escherichia coli BL21 (DE3). The best linker ligating the two enzymes was a flexible peptide containing 12 residues (GSAGSAAGSGEF). At 55 °C and pH 8.0, and as with the addition of 1 mmol L-1 Mn2+ , the CSCA-linkerE-DAE fusion enzyme obtained through high cell-density cultivation displayed a maximal exo-inulinase activity of 21.8 U mg-1 and resulted in a yield of 6.3 g L-1 d-allulose and 39.2 g L-1 d-fructose using 60 g L-1 inulin as the raw material. Catechol-modified alginate with titanium ions (Alg(Ti)PDA) was found to be a promising immobilization material for the fusion enzyme. After conversion for 8 days, the Alg(Ti)PDA-immobilized CSCA-linkerE-DAE (8 U g-1 ) completed 24 reaction cycles and retained over 80% of its original activity. Each reaction obtained an average of 19.8 g L-1 d-allulose and 32.7 g L-1 D-fructose from 60 g L-1 inulin. CONCLUSION: This study shed light on a feasible and cost-effective approach for the production of syrup containing d-allulose and D-fructose with inulin as the raw material via the use of a CSCA and DAE fusion enzyme. This syrup is of added value as a functional sweetener. © 2020 Society of Chemical Industry.

Assessment of renal function in living kidney donors before and after nephrectomy: A Japanese prospective, observational cohort study.

OBJECTIVE: To investigate the utility of estimated glomerular filtration rate for assessing kidney function in living kidney donors before and after nephrectomy. METHODS: A total of 101 donors underwent inulin clearance measurements before and 1 year after nephrectomy. The mean of three inulin clearance values was used as the measured glomerular filtration rate. Estimated glomerular filtration rate based on serum creatinine and cystatin C levels was calculated using the Japanese estimated glomerular filtration rate equation, Chronic Kidney Disease Epidemiology Collaboration formula and new full age spectrum equation. Age-adjusted chronic kidney disease was defined as glomerular filtration rate <75 mL/min/1.73m2 for donors aged <40 years, <60 mL/min/1.73m2 for donors aged 40-65 years and <45 mL/min/1.73m2 for donors aged >65 years. RESULTS: The postoperative measured glomerular filtration rate <60 mL/min/1.73m2 and age-adjusted chronic kidney disease rate were 36.0% and 27.0%, respectively. In younger donors (aged <50 years), postoperative measured glomerular filtration rate <60 mL/min/1.73m2 and age-adjusted chronic kidney disease rates were 5.3% and 26.3%, respectively. In older donors (aged >70 years), postoperative measured glomerular filtration rate <60 mL/min/1.73m2 and age-adjusted chronic kidney disease rates were 75.0% and 33.3%, respectively. Donor age and measured glomerular filtration rate were significant predictors of postoperative measured glomerular filtration rate. The Japanese estimated glomerular filtration rate equation based on creatinine and cystatin C showed the strongest correlation with measured glomerular filtration rate. However, the Japanese estimated glomerular filtration rate equation based on creatinine overestimated the prevalence of measured glomerular filtration rate <60 mL/min/1.73m2 , whereas the Japanese estimated glomerular filtration rate based on cystatin C underestimated it. CONCLUSIONS: Aged donors might have an increased risk of lower glomerular filtration rate after donor nephrectomy; post-surgery, long-term monitoring of renal function is recommended. Measurement of glomerular filtration rate should be carried out for donors, especially pre-surgery. A more precise glomerular filtration rate equation is required in the future.

Statistical optimization of process parameters for inulinase production from Tithonia weed by Arthrobacter mysorens strain no.1.

Tithonia rotundifolia is an easily available and abundant inulin rich weed reported to be competitive and allelopathic. This weed inulin is hydrolyzed by inulinase into fructose. Response surface methodology was employed to optimize culture conditions for the inulinase production from Arthrobacter mysorens strain no.1 isolated from rhizospheric area of Tithonia weed. Initially, Plackett- Burman design was used for screening 11 nutritional parameters for inulinase production including inulin containing weeds as cost effective substrate. The experiment shows that amongst the 11 parameters studied, K2HPO4, Inulin, Agave sisalana extract and Tithonia rotundifolia were the most significant variables for inulinase production. Quantitative effects of these 4 factors were further investigated using Box Behnken design. The medium having 0.27% K2HPO4, 2.54% Inulin, 6.57% Agave sisalana extract and 7.27% Tithonia rotundifolia extract were found to be optimum for maximum inulinase production. The optimization strategies used showed 2.12 fold increase in inulinase yield (1669.45 EU/ml) compared to non-optimized medium (787 EU/ml). Fructose produced by the action of inulinase was further confirmed by spectrophotometer, osazone, HPTLC and FTIR methods. Thus Tithonia rotundifolia can be used as an eco-friendly, economically feasible and promising alternative substrate for commercial inulinase production yielding fructose from Arthrobacter mysorens strain no.1.

Enhanced Inulin Saccharification by Self-Produced Inulinase from a Newly Isolated Penicillium sp. and its Application in D-Lactic Acid Production.

In order to find an alternative for commercial inulinase, a strain XL01 identified as Penicillium sp. was screened for inulinase production. The broth after cultivated was centrifuged, filtered, and used as crude enzyme for the following saccharification. At pH 5.0 and 50 °C, the crude enzyme released 84.9 g/L fructose and 20.7 g/L glucose from 120 g/L inulin in 72 h. In addition, simultaneous saccharification and fermentation of chicory flour for D-lactic acid production was carried out using the self-produced crude inulinase and Lactobacillus bulgaricus CGMCC 1.6970. A high D-lactic acid titer and productivity of 122.0 g/L and 1.69 g/(L h) was achieved from 120 g/L chicory flour in 72 h. The simplicity for inulinase production and the high efficiency for D-lactic acid fermentation provide a perspective and profitable industrial biotechnology for utilization of the inulin-rich biomass.

Assessment of probiotic, antifungal and cholesterol lowering properties of Pediococcus pentosaceus KCC-23 isolated from Italian ryegrass.

BACKGROUND: Lactic acid bacteria (LAB) are important for the processing of various food products. Although genetically modified organisms have contributed to improvements in various food products, there are some limitations. Thus, the discovery of wild strains from natural sources must be considered as the most suitable approach for identifying new LAB. Therefore, we planned to isolate and characterise the LAB from Italian ryegrass forage and evaluate their biological potential. RESULTS: A total of 28 strains were isolated and screened for their anti-fungal and probiotic properties. A single strain was selected due to its antifungal and probiotic efficiency. The strain was identified as Pediococcus pentosaceus KCC-23. The strain KCC-23 showed effective inhibition against Aspergillus fumigatus, Pencillium chrysogenum, Pencillium roqueforti, Botrytis elliptica and Fusarium oxysporum. Further, it survived low pH, and the presence of bile salts and gastric juice. It exhibited significant aggregation and hydrophobicity properties. The KCC-23 effectively assimilated cholesterol and had the ability to utilise pre-biotics such as raffinose and inulin. Finally, KCC-23 exhibited significant free radical scavenging activity. CONCLUSION: P. pentosaceus KCC-23 showed effective anti-fungal, probiotic and anti-oxidant properties and would be a promising isolate for exploitation in the formulation of food for ruminants and humans.

Optimization of inulinase production from low cost substrates using Plackett-Burman and Taguchi methods.

Four marine-derived fungal isolates were screened for the production of inulinase enzyme from low cost substrates under solid state fermentation (SSF), one of them identified as Aspergillus terreus showed the highest inulinase activity using artichoke leaves as a solid substrate. Sequential optimization strategy, based on statistical experimental designs was employed to optimize the composition of the medium, including Plackett-Burman and Taguchi's (L9 3(4)) orthogonal array designs. Under the optimized conditions, inulinase activity (21.058 U/gds) reached the predicted maximum activity derived from the taguchi methodology, which increased about 4.79-folds the initial production medium. Fructose was produced, as an end product of inulin hydrolysis proving that the enzyme produced was exoinulinase. The marine-derived A. terreus is suggested as a new potential candidate for industrial enzymatic production of fructose from low cost substrate containing inulin as an economic source.

Techniques for the in vivo assessment of cardio-renal function in zebrafish (Danio rerio) larvae.

Zebrafish, a well-established vertebrate model, offer unique advantages for assessing renal function and physiology. Assays determining renal glomerular function based on cardiovascular erythrocyte flow and reduction of injected FITC-inulin were developed, each validated using the nephrotoxin gentamicin. Bland­Atlman analysis showed a strong association between measurements of the rate of inulin excretion and that of fluorescent reduction from the arterial vasculature. Reduced renal clearance of inulin, resulting from gentamicin or NaCl loading, was concurrent with reduced erythrocyte velocity, and yolk sac and pericardium oedema. These techniques, assessing pronephric function, highlight the potential for in vivo physiological study in this genetically tractable model.

Microbial production of 2,3-butanediol from Jerusalem artichoke tubers by Klebsiella pneumoniae.

2,3-Butanediol is one of the promising bulk chemicals with wide applications. Its fermentative production has attracted great interest due to the high end concentration. However, large-scale production of 2,3-butanediol requires low-cost substrate and efficient fermentation process. In the present study, 2,3-butanediol production by Klebsiella pneumoniae from Jerusalem artichoke tubers was successfully performed, and various technologies, including separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF), were investigated. The concentration of target products reached 81.59 and 91.63 g/l, respectively after 40 h in batch and fed-batch SSF processes. Comparing with fed-batch SHF, the fed-batch SSF provided 30.3% higher concentration and 83.2% higher productivity of target products. The results showed that Jerusalem artichoke tuber is a favorable substrate for 2,3-butanediol production, and the application of fed-batch SSF for its conversion can result in a more cost-effective process.

Assessment of dietary fiber fermentation: effect of Lactobacillus reuteri and reproducibility of short-chain fatty acid concentrations.

This investigation had two aims: (i) to determine the reproducibility of SCFA production of two fibers: wheat dextrin and inulin, in two separate in vitro batch fermentation systems, and (ii) to determine if the addition Lactobacillus reuteri, a probiotic bacterium, enhanced the fermentation of wheat dextrin, inulin, and psyllium using in vitro batch fermentation. Samples were removed at 0, 4, 8, 12, and 24 h. SCFAs were measured by GC. L. reuteri improved inulin's fermentation profile by reducing the total SCFA peak at 4 h and enhancing fermentation at 8 and 12 h. Wheat dextrin and psyllium were largely unaffected. Wheat dextrin's total SCFA and propionate production curves were steady and replicable, but concentration values varied between fermentations. Partially hydrolyzed guar gum (PHGG) and wheat dextrin had similar fermentation patterns from 0-8 h, but PHGG plateaued at 8 h for all measures. Psyllium produced peak SCFA concentrations at 8 h, similar to inulin. L. reuteri could be combined with inulin for enhancing fermentation, but it does not improve wheat dextrin or psyllium fermentation. Wheat dextrin will likely produce similar physiological within a group of individuals due to the reproducibility of fermentation.

Glomerular filtration rate assessment in individuals after orthotopic liver transplantation based on serum cystatin C levels.

Individuals after orthotopic liver transplantation (OLT) often show renal dysfunction, which may substantially affect the post-OLT course. Renal function after OLT is commonly assessed by means of serum creatinine (S(cr)) concentration or renal creatinine clearance (C(cr)). A glomerular filtration rate (GFR) estimate based on S(cr) level is not accurate enough because even a more marked decrease in GFR need not be associated with an increase in S(cr) level, especially in jaundiced patients. The study intends to try to estimate GFR in individuals after OLT by means of determining serum cystatin C (S(cyst)) concentrations. In 58 individuals (mean age, 49 +/- 7 years; 31 men, 27 women) at various intervals from OLT (mean, 14 +/- 10 months), GFR was estimated by using simultaneous determinations of S(cyst), S(cr), C(cr), and renal inulin clearance (C(in)). In most subjects (91.3%), C(in) was decreased to less than the lower limit of normal (80 mL/min/1.73 m(2)). A significant correlation (r = 0.70; P <.001) was found between 1/S(cyst) and C(in). Receiver operating characteristic analysis was performed on S(cyst) and S(cr) using a C(in) cutoff value of 80 mL/min/1.73 m(2). The area under the curve for S(cyst) was 0.912 +/- 0.044, and that for S(cr), 0.899 +/- 0.049. There was no statistically significant difference between these values. The sensitivity for a S(cyst) level of 1.20 mg/L (upper limit of normal value) to detect a decrease in GFR (measured as C(in)) below the lower limit of normal (80 mL/min/1.73 m(2)) was 96.1%. The sensitivity of S(cyst) level was significantly greater (P <.01) than the sensitivity of S(cr) level for men and at borderline significance for women (P =.05). Findings support the assumption that a S(cyst) level less than 1.2 mg/L indicates with a high degree of probability (P <.001) that GFR is not decreased to less than the normal limit. S(cyst) assessment in individuals after OLT could be proposed as a confirmatory test of a decrease in GFR in individuals with normal S(cr) levels.

Assessment of the fructanolytic activities in the rumen bacterium Treponema saccharophilum strain S.

AIMS: To characterize the fructose polymer degrading enzymes of rumen bacterium Treponema saccharophilum strain S. METHODS AND RESULTS: Conventional methods were used to examine bacterial growth and enzyme activities. Electrophoretic zymogram under native conditions, and thin layer chromatography, were applied to identify and characterize the enzymes. Treponema saccharophilum utilized Timothy grass fructan, inulin and sucrose but not free fructose. Timothy grass fructan was degraded at a significantly higher rate than sucrose and inulin. Two fructanolytic enzymes were found in the soluble, and one in the membrane fraction of bacterial cell extract. The first degraded each mentioned carbohydrate to monosaccharides. The second released oligosaccharides only from Timothy grass fructan. CONCLUSIONS: The bacterium T. saccharophilum strain S is capable of synthesizing non-specific beta-fructofuranosidases and 2,6-beta-D-fructan fructanohydrolase. The enzymes are of constitutive character. SIGNIFICANCE AND IMPACT OF THE STUDY: It has been stated for the first time that the 2,6-beta-D-fructan fructanohydrolase is synthesized by the rumen bacterium T. saccharophilum. This organism appears to be responsible for grass fructan degradation in the rumen.

Selecting the best renal function tests. A meta-analysis of diagnostic studies.

A literature review compared the results of different kidney function tests using the inulin clearance or similar methods as a gold standard. The methodological strength of the available studies was weak. For the time being, the calculated creatinine clearance using formula, is a first choice, especially in general practice.

Assessment of renal function by inulin clearance: comparison with creatinine clearance as determined by enzymatic methods.

We compared creatinine clearances determined by enzymatic (Kodak Ektachem 700 single-slide, Boehringer Mannheim creatinine PAP) and nonenzymatic (Jaffé, HPLC) methods with glomerular filtration rate measured by inulin clearance in patients with varying degrees of renal function. The Kodak enzymatic assay gave values for creatinine 2 to 3 mg/L higher than the other methods. This resulted in significantly lower creatinine clearances than inulin clearances and creatinine clearances determined by the other methods. However, correlations between all methods for serum and urinary creatinine values and clearances were good. To avoid between assay (enzymatic vs nonenzymatic) discrepancies, manufacturers should agree to an acceptable standard of calibration under the usual conditions used with patients.

Serial assessment of glomerular filtration rate in lupus nephropathy.

In patients with lupus nephropathy (LN), previous studies have shown that creatinine clearance (CCr) overestimates true glomerular filtration rate as measured by inulin clearance (CIn), and that among patients the degree of overestimation is highly variable. We sought to determine whether the discrepancy between CCr and CIn remains constant over time (months, years) in each individual patient, and therefore whether serial measurements of CCr reliably reflect the direction and magnitude of change in CIn. Twenty-five patients with LN underwent simultaneous determinations of CCr and CIn performed two to four (mean 3.3) times over three years. In a given patient, it was found that the ratio of CCr/CIn changed substantially over time (mean SD 0.16 with 95% confidence interval of 0.12 to 0.20). Thus, in about 32% of cases the ratio of CCr/CIn will vary more than +/- 16% from a previously measured value of CCr/CIn. Patients with both high and low values of CIn showed similar variability in CCR/CIn over time. Variability in CCr/CIn was found regardless of whether CIn was increasing, decreasing, or constant over time. In nearly one-half of all measurements of CCr, the corresponding change in CIn was directionally discordant. Iothalamate and technetium-DTPA renal clearances correlated highly with CIn (R2 = 0.99). We conclude that the discrepancy between CCr and CIn can vary greatly over time in an individual patient. Consequently, serial CCr does not accurately measure the direction or magnitude of change in glomerular filtration rate in lupus nephropathy.

Simple assessment of the efficacy of peritoneal transport in continuous ambulatory peritoneal dialysis patients.

The efficacy of peritoneal transport was assessed in 13 permeability studies in 11 continuous ambulatory peritoneal dialysis (CAPD) patients. During each study the in situ intraperitoneal volume was measured as well as the dialysate and plasma concentrations of various solutes with a molecular weight range from 60 to 5,500. As clearance estimations are unsuitable for the purpose of permeability studies, mass transfer area coefficients were used. By applying a simple mathematical model assuming first-order kinetics, these coefficients were calculated for urea, lactate, creatinine, glucose, kanamycin, and inulin. The accuracy of the calculations is indicated by their r values. After pooling these correlation coefficients, the mean approached 1.00 for all solutes with high confidence limits, indicating the usefulness of the model. A further simplification was tested using only an initial- and end-dialysate sample and two blood samples, without the measurement of the in situ intraperitoneal volume. Except for inulin the results of this simplification correlated well with the results described above. The reproducibility of the simplified mass transfer area coefficient calculations was investigated on 15 occasions in 3 other CAPD patients. The coefficients of variation of low molecular weight solutes varied between 15 and 20%. It is concluded that mass transfer area coefficient estimations using the latter method can be performed in any CAPD patient and probably yield sufficient information to establish the efficacy of the membrane transport mechanism during clinical follow-up.

Assessment of compensatory renal function and [125I]iothalamate clearance in sheep.

Compensatory renal function and sodium [125I]iothalamate clearance were studied in 4 adult Merino ewes. Renal clearances of inulin (CIN), iothalamate (CIOT), and p-aminohippurate (CPAH) were used to measure renal function before and 7 and 170 days after 50% and 75% reductions in renal mass. Renal mass reduction to 50% of normal was achieved by ligation of the right renal artery, and to 25% of normal by ligating the right renal artery and branches of the left renal artery. Renal function (CIN and CPAH) after 50% renal mass reduction was 78% of normal on postligation day (PLD) 7 and 64% of normal on PLD 170. This decrease in renal function between PLD 7 and 170 was significant (P less than 0.05). Renal function (CIN and CPAH) after 75% renal mass reduction was 55% of normal on PLD 7 and 48% on PLD 170; however, this decrease in renal function between PLD 7 and 170 was not significant. Results indicated that, shortly after 50% renal mass reduction by renal arterial ligation, renal function in ewes was greater than that previously reported after unilateral nephrectomy and that a further decrease in renal function developed within 6 months after ligation. The mean CIOT in healthy ewes was 2.46 ml/min/kg of body weight. In healthy ewes and in ewes after renal mass reduction, regression of the simultaneous CIOT and CIN was CIOT = 0.276 + 1.05 CIN. The CIOT was higher than the corresponding CIN, and remained significantly (P less than 0.01) and consistently higher. Therefore, iothalamate can be used to estimate glomerular filtration rate in sheep, although a correction factor is required.

A comparison of micropuncture and lithium clearance methods in the assessment of renal tubular function in rats with diabetes insipidus.

The lithium clearance technique has been proposed as a non-invasive method whereby fluid delivery from the pars recta and pars convoluta of proximal tubules can be measured as CLi and CIN [0.78 CLi/CIN + 0.22], respectively [12], CLi being the clearance of lithium and CIN that of inulin. In the present study, fluid delivery from proximal tubules was estimated simultaneously by micropuncture and lithium clearance techniques in anaesthetized Brattleboro rats with diabetes insipidus, under control conditions and following chronic treatment with hydrochlorothiazide. Absolute deliveries from the proximal convoluted tubules as determined by the micropuncture and lithium clearance methods were 437 and 427 microliter/min, respectively, in untreated animals and 348 and 355 microliter/min, respectively, in thiazide-treated animals. The individual results obtained by the two methods showed a high degree of correlation (r = 0.85, P less than 0.001). In untreated Brattleboro rats, proximal fluid delivery as estimated by both the micropuncture and lithium clearance techniques showed significant (P less than 0.001) correlations with urine flow rate. These results provide further evidence for the acceptance of lithium clearance as a valid estimate of proximal tubular fluid delivery.