Purine Base Concentrations Research Articles

Purine metabolism state in patients with type 1 diabetes mellitus

The aim of the study. To carry out a comprehensive analysis of the purine metabolism (PM) state and assess its features in patients with type 1 diabetes mellitus (DM). Materials and methods. 181 patients with type 1 DM were examined (94 women and 87 men) aged 42.5 ± 12.1 years. Indicators of the carbohydrate metabolism state, creatinine (Cr) concentration by the method of Popper, purine bases (PBs) and the activity of xanthine oxidase (XO) by a photometric method, uric acid (UA) by a colorimetric method were determined in fasting venous blood. UA excretion was detected by the colorimetric method, Cr by an enzymatic method. Renal UA clearance (RCUA), fractional UA clearance (FCUA), total UA tubular reabsorption (TTRUA) and hypoxanthine-guanine-phosphoribosyltransferase (HGPRT) activity were calculated. The comparison group, representative in terms of age and sex, included 25 healthy volunteers. Results. PM changes in patients with type 1 DM were orientated towards an excessively increased catabolism and insufficient reutilization of PBs. The structure of the detected disorders was as follows: hyperuricemia (HU) (13.8 %), enhanced RCUA (42.8 %), increased XO activity (35.6 %) and inhibition of HGPRT activity (53.3 %). In about 56 % of the subjects, high concentrations of PBs were found, and HU was diagnosed only in every seventh subject. It was identified that PB concentrations were negatively correlated with the level of XO activity. RCUA and XO activity levels were revealed to be of the greatest informational value for assessing the PM state in patients with type 1 DM. RCUA was significantly associated with the level of UA excretion and HGPRT activity. Relationships of RCUA and FCUA with HbAc1 levels were established. The higher the level of HbAc1, the greater the clearance of RCUA was, especially FCUA, which led to a significant decrease in TTRUA. Conclusions. PM in type 1 DM is characterized by a high intensity, which is realized due to a decrease in anabolism, increased oxidation and suppression of PB reutilization. Uricemia inadequately reflects the level of UA production in patients with type 1 DM. The severity of PM disorders in type 1 DM patients is associated with the carbohydrate metabolism compensation state.

Evaluation of 15N and purine bases as microbial markers to estimate ruminal bacterial nitrogen outflow in dairy cows

The present study aimed to evaluate 15N and purine bases (PB) as markers for ruminal bacterial nitrogen (N) outflow in dairy cows. Additionally, the effects of calcium salts of fatty acids (CSFA) in diets with different levels of rumen degradable protein (RDP) on PB omasal flow, urinary recovery of purine derivatives (PD), and endogenous excretion of PD were evaluated. Eight ruminal cannulated Holstein cows, with 207 ± 23.6 (mean ± SD) days in milk, 618 ± 39.0 kg of body weight, and 33.4 ± 2.45 kg/d of milk yield were used in a replicated 4 × 4 Latin square design. Four treatments were arranged in a 2 × 2 factorial design to evaluate the following factors: CSFA addition [-CS: basal diet without CSFA; and + CS: with 33.2 g/kg dry matter (DM) of CSFA]; and the level of RDP (LRDP: low RDP, with 98 g/kg DM of RDP; and HRDP: high RDP, with 110 g/kg DM of RDP). Particle-associated bacteria (PAB) showed lower (P ≤ 0.008) 15N enrichment and purine N to total bacterial N ratio than liquid associated bacteria (LAB). Increasing RDP concentration reduced (P = 0.007) 15N enrichment and tended (P ≤ 0.095) to reduce PB concentration in bacterial pellets. Purine bases overestimated (P ≤ 0.002) LAB-NAN and total microbial flow but, regarding PAB-NAN flow, there was a trend of interaction (P = 0.067) between marker and fat supplementation effects. In the absence of fat supplementation, PB underestimated (P ≤ 0.05) the flow of PAB-NAN compared to 15N and failed to detect the negative effect of fat supplementation evidenced by 15N labeling. Omasal PB flow tended to increase (P = 0.061) in low protein degradability diets, but PD excretion was constant (322 ± 5,9 mmol/d, mean ± SE) regardless of diet and, as a result, urinary recovery of PB decreased (P = 0.007). Daily excretion of creatinine and endogenous PD was also unaffected by the diet and averaged 0.262 mmol/kg BW and 0.640 mmol/kg BW0.75, respectively. In conclusion, the response of PB and its urinary derivatives to variation in diet composition compared to 15N reveals their weaknesses in the estimation of microbial synthesis.

In vitro rumen fermentation, microbial protein synthesis and composition of microbial community of total mixed rations replacing maize silage with red clover silage.

This study aimed to evaluate in vitro fermentation characteristics, microbial protein synthesis and microbial community composition when replacing maize silage (MS) with red clover silage (RCS) in total mixed rations (TMR) of dairy cows. Treatments included TMR containing forage (MS and RCS) and concentrates (0.75:0.25) with targeted proportions (dry matter (DM) basis) of RCS in TMR of 0.15 (RCS15 ), 0.30 (RCS30 ), 0.45 (RCS45 ), and 0.60 (RCS60 ), in substitution of MS. Samples of the TMR were incubated using the in vitro Ankom RF technique with a mixture of rumen fluid and buffer solution (1:2 v/v) for 8 and 24hr. Gas production and total short-chain fatty acids concentration did not differ between diets, whereas ammonia-nitrogen concentration increased with increasing level of RCS. Acetate proportion was not affected by RCS level, but propionate showed a linear increase with increasing level of RCS at the expenses of butyrate. Branched fatty acids proportions linearly declined, reflecting a reduced deamination of true protein. Gene copy numbers of protozoa linearly decreased with increasing RCS levels, while total numbers of bacteria and methanogens were not affected by diet. The amylolytic bacteria Ruminobacter amylophillus and Prevotella bryantii showed evidence to increase with higher RCS levels after 8hr and 24hr, respectively, whereas no effects of diet where observed for the fibrolytic bacteria Ruminococcus albus, Ruminococcus flavefaciens and Fibrobacter succinogenes. Concentrations of purine bases, and total N production in liquid-associated microbes declined with increasing RCS levels, suggesting a negative impact of this feed on microbial growth. The findings of this study suggest that in general, microbial protein synthesis might be impaired by the substitution of MS by RCS, therefore caution should be taken when formulating diets for dairy cows using high levels of RCS as ingredient.

Postruminal digestion of starch infused into the abomasum of heifers with or without exogenous amylase administration.

The effect of an exogenous amylase on postruminal digestion of starch infused into the abomasum of cattle was studied. Four rumen-cannulated heifers were fed 5.5 kg DM/d of a diet without starch, and assigned randomly to a crossover design. The experiment consisted of 2 periods lasting 23 d each with 10 d for adaptation to the diet followed by 13 d of abomasal infusion and sample collection. During the first 3 d of each infusion phase, isotonic saline solution was infused (1 liter/h) for measurement of baseline values in feces, followed by daily infusions of 880 g DM corn starch (1 kg/10 liters of water) without or with the addition of 2% of amylase. Titanium dioxide (10 g/d) was ruminally administered for estimation of fecal excretion. Digestion of starch in small intestine was calculated as the difference between the amounts of infused starch, disappeared from hindgut and fecal excretion. The apparent disappearance of starch from the hindgut was estimated based on the increment of microbial nitrogen (N) excretion due to starch infusion (1 g microbial N/100 g fermented starch) compared to baseline values. The concentration of purine bases in feces was used to estimate excretion of microbial N. Microbial N excretion increased with starch infusion (P < 0.05) but was not influenced by amylase (P = 0.81). Starch disappearance from the small intestine was not improved by amylase (P = 0.78) and averaged 85%. Amylase affected neither blood concentration of glucose (P = 0.80) nor of insulin (P = 0.26), but glucagon was lower without (P < 0.0001) than with amylase. The infusion of starch increased fecal excretion of total VFA (acetate, propionate, and butyrate) by 53% (P < 0.05), which indicates increased carbohydrate fermentation in the hindgut and incomplete digestion of starch in the small intestine. However, the excretion of total VFA was not affected by amylase (P = 0.66). Lactate excretion was higher at the second day of starch infusion (P < 0.05) without than with amylase, which suggests lower flow of starch from the small intestine to the hindgut due to a possible effect of amylase addition in animals not adapted to starch digestion. However, lactate excretion returned near to baseline values within 2 d, which was probably due to increase of lactate-utilizing bacteria and the adaptation of the microbial population in the hindgut. Further studies with higher starch levels and addition of amylase are recommended.

Effects of synchronicity of carbohydrate and protein degradation on rumen fermentation characteristics and microbial protein synthesis.

A series of in vitro studies were carried out to determine i) the effects of enzyme and formaldehyde treatment on the degradation characteristics of carbohydrate and protein sources and on the synchronicity of these processes, and ii) the effects of synchronizing carbohydrate and protein supply on rumen fermentation and microbial protein synthesis (MPS) in in vitro experiments. Untreated corn (C) and enzyme-treated corn (EC) were combined with soy bean meal with (ES) and without (S) enzyme treatment or formaldehyde treatment (FS). Six experimental feeds (CS, CES, CFS, ECS, ECES and ECFS) with different synchrony indices were prepared. Highly synchronous diets had the greatest dry matter (DM) digestibility when untreated corn was used. However, the degree of synchronicity did not influence DM digestibility when EC was mixed with various soybean meals. At time points of 12 h and 24 h of incubation, EC-containing diets showed lower ammonia-N concentrations than those of C-containing diets, irrespective of the degree of synchronicity, indicating that more efficient utilization of ammonia-N for MPS was achieved by ruminal microorganisms when EC was offered as a carbohydrate source. Within C-containing treatments, the purine base concentration increased as the diets were more synchronized. This effect was not observed when EC was offered. There were significant effects on VFA concentration of both C and S treatments and their interactions. Similar to purine concentrations, total VFA production and individual VFA concentration in the groups containing EC as an energy source was higher than those of other groups (CS, CES and CFS). The results of the present study suggested that the availability of energy or the protein source are the most limiting factors for rumen fermentation and MPS, rather than the degree of synchronicity.

METHANOGENESIS IN RABBIT CAECUM AS AFFECTED BY THE FERMENTATION PATTERN: IN VITRO AND IN VIVO MEASUREMENTS

Methane formation and caecal fermentation patterns were studied in vivo and in vitro in 16 white New Zealand rabbits (70-80 d and 2.27 ± 0.064 kg) allocated to four diets formulated to have a similar neutral detergent fibre (33.8±0.53%) and protein (17.7±0.33%) content, with two different fibre sources (alfalfa hay, AH or sugar beet pulp, SP) and starch (wheat or maize). Animals received the diet for 16 to 20 days before methane production was measured in vivo in a respiratory chamber. Animals were subsequently slaughtered at approximately 9:00 and caecal contents were sampled and used as inoculum for in vitro incubations to determine gas and methane production. Volatile fatty acid (VFA) and purine base (PB) concentrations were determined from both caecal content and incubation medium after 6 h. Total VFA concentration in caecal content decreased (P&lt;0.05) in rabbits fed AH-maize diet compared with rabbits fed AH-wheat and SP-maize diets (37.7 vs. 59.6 mM), with those fed SP-wheat showing an intermediate value (53.0 mM). Fermentation pattern was affected when maize was the source of starch compared to wheat, with lower acetate (0.72 vs. 0.79; P&lt;0.01) and higher butyrate (0.19 vs. 0.14; P&lt;0.001) molar proportions. Fermentation in vivo vs. in vitro showed some differences (molar proportions of acetate, 0.76 vs. 0.73, P&lt;0.001, and propionate, 0.069 vs. 0.091, P&lt;0.001, in vivo and in vitro, respectively), probably due to differences in pH (6.0 vs. 6.7 in vivo and in vitro; P&lt;0.001). Only 2 out of 16 rabbits produced a substantial volume of methane in vivo (on average, 12.6 ml/BW0.75/d or 0.56 mmol/BW0.75/d), showing a high inter-individual variability that hindered comparison of treatment differences. In contrast, methane was detected in vitro in all cases and volumes were more homogenous, a higher formation (P&lt;0.05) being observed with maize compared to wheat. A similar effect was shown in total gas production. The low methane production and H2 recovery suggest the importance of H2 disposal mechanisms other than methanogenesis, such as reductive acetogenesis. PB concentration in caecal content and the incubation medium, as an index of microbial concentration, was highest when SP was added with maize (P&lt;0.05).

Technical note: The persistence of microbial-specific DNA sequences through gastric digestion in lambs and their potential use as microbial markers1

Two groups of 5 lambs were euthanized at the weaning (T45) and fattening stages (T90) to evaluate the use of microbial ribosomal DNA (rDNA) sequences as potential microbial markers in relation to purine bases (PB) as a conventional marker. Both microbial markers originated similar microbial N concentrations (mg/g of DM), although T45 showed decreased values compared with the T90 group when either PB or rDNA were considered (P = 0.02). The survival of microbial rDNA was determined in 3 digestive sites (omasum, abomasum, and duodenum), but no substantial differences were observed, indicating that rDNA maintains the molecular stability along the sampling sites analyzed. Contrarily PB concentration increased successively along the digestive tract (P < 0.05), likely as a consequence of the endogenous PB secretion. Undegraded milk PB may also explain the overestimation of the microbial N concentration (2.8 times greater) using PB than rDNA sequences. Abomasum was the sampling site where the best agreement between PB and rDNA estimations was observed. Protozoal N concentration was irrelevant in T45 animals, although substantial in T90 lambs (18% of microbial N). In conclusion, bacterial 16S and protozoal 18S rDNA sequences may persist through the gastric digestive tract and their utilization as a highly specific microbial marker should not be neglected.

Effects of extracellular purines on cytotoxicity of methotrexate

Nucleoside and base modulation of the cytotoxicity of nucleic acid and folate antimetabolite drugs has been widely discussed. Many investigators have observed reduced toxicity due to circumvention of drug-induced inhibition of de novo purine and pyrimidine synthesis. However, exogenous purine nucleosides and bases may also enhance the cytotoxicity of even moderate concentrations of antifolate drugs (MTX and PTX) which inhibit dihydrofolate reductase. In this study, the effects of nucleosides in the medium on the cytotoxicity and deoxyribonucleoside triphosphate pools after brief exposure of cultured cells to methotrexate have been studied in cultured L1210 murine leukaemia cells. Cell viability was determined by trypan blue exclusion assay. Colony formation was assessed by microtitration cloning assay. The deoxyribonucleotides were measured by a modification of the DNA polymerase assay. Purines were extracted with trioctylamine and 1,1,2-trichlorotrifluoroethane buffer and concentrations of purine bases were determined by HPLC. Subculture of drug-treated cells in fresh medium containing 10% FCS led to greater toxicity than sub culture in 'conditioned' medium, i.e. fresh medium in which logarithmically growing cells had been cultured for 24 h before separation. Cells resuspended in fresh medium had increased dATP and sustained inhibition of dTTP levels, while cells subcultured in 'conditioned' medium had no elevation of dATP. Hypoxanthine concentration determined by HPLC in 'conditioned' medium was 0.9 microM compared to 6.7 microM in fresh medium. Resuspension of drug-treated cells in conditioned medium supplemented with 10 or 100 microM HX enhanced cytotoxicity and increased the dATP levels. These results add further evidence that purines present in normal culture conditions are important determinants of methotrexate cytotoxicity. Elevation of dATP levels after methotrexate treatment is an important modulator of cytotoxicity.

In vitro ruminal fermentation of Pennisetum purpureum CT-115 supplemented with four tropical browse legume species

An in vitro gas production experiment was completed to determine effects of four tropical browse legumes ( i.e., Acacia cornigera, AC; Albizia lebbekoides, AL; Enterolobium cyclocarpum, EC; Leucaena leucocephala, LL) at 0.15 or 0.30 of total substrate on microbial fermentation of the tropical grass Pennisetum purpureum CT-115 (PP). Two incubation runs with four bottles per treatment were completed, using two bottles for gas production measurements up to 96 h and two bottles for microbial purine bases (PB) concentration analysis after 24 h. Inclusion of AC, EC and LL linearly increased ( P<0.05) gas production from PP alone at an initial stage of fermentation (5 h) because of their higher fermentation rate versus PP. However, whereas the linear increase in gas production was maintained at mid fermentation ( i.e., 24 h) with AC, there was no effect with LL and a linear reduction in gas volume at 24 and 96 h with EC, possibly because of its saponin content. Gas production with AL was linearly reduced at 24 and 96 h ( P<0.01). There were no differences in microbial concentrations, either estimated from a partitioning factor or PB analysis, but PB concentration in the solid residue increased three-fold when LL were included. Differences among legume species in chemical composition, and in the nature of their secondary compounds, are responsible for the fermentation pattern of mixed substrates.

Rumen fermentation, microbial protein synthesis, and nutrient flow to the omasum in cattle offered corn silage, grass silage, or whole-crop wheat1

The objectives of this study were to determine the relative effect of feeding corn silage (CS), fermented whole-crop wheat (FWCW), and urea-treated processed whole-crop wheat (UPWCW) compared with grass silage (GS), each supplemented with concentrates, on forage intake, ruminal fermentation, microbial protein synthesis, some plasma metabolites, and ruminal and total tract digestibility in cattle. Four ruminally fistulated steers with a mean BW of 509 kg (SD 6.3) were used in a 4 x 4 Latin square-designed experiment with each period lasting 21 d. The omasal sampling technique in combination with a triple marker method was used to measure nutrient flows to the omasum with Co-EDTA, Yb acetate, and indigestible NDF as liquid, small particle, and large particle phase markers, respectively. Microbial N flow was assessed from purine base concentrations. Steers fed CS, FWCW, and UPWCW consumed 2.7, 2.4, and 2.6 kg/d more (P < 0.05) forage and total DMI, respectively, than those fed GS-based diets. Rumen pH (P = 0.07) and lactic acid (P = 0.11) concentration did not differ between the forages. Rumen concentration of NH(3)-N was greatest for UPWCW and least for CS (P < 0.001). Total VFA concentrations were greater (P < 0.05) for CS than GS and UPWCW, with FWCW being intermediate. Acetate-to-propionate ratio (P < 0.05) was greater (P < 0.05) for UPWCW than the other forages, which did not differ. Apparent ruminal digestion of OM (P < 0.05) was less for CS, FWCW, and UPWCW than GS. Ruminal NDF digestibility was greater (P < 0.01) for GS than the other forages, which did not differ (P > or = 0.06). Total tract NDF digestibility was less (P < 0.05) for UPWCW than the other forages, with GS being greatest and CS and FWCW being intermediate. Starch intake was less (P < 0.001) for GS than the other forages, but there was no effect of forage on omasal starch flow (P = 0.23) or ruminal digestibility (P = 0.88). Flow of non-NH(3)-N and microbial N was greater (P < 0.05) for CS, FWCW, and UPWCW than GS. Efficiency of microbial N synthesis was greater (P < 0.05) for FWCW than GS and CS, with UPWCW being intermediate. Plasma beta-hydroxybutyrate concentrations were greatest with CS and least for GS (P < 0.001), whereas concentrations of plasma urea were least for CS and greatest for UPWCW (P < 0.001). In conclusion, feeding alternative forages to GS can significantly increase feed DMI and alter rumen fermentation and site of nutrient digestion when offered to cattle supplemented with 3 kg of concentrate daily.

Effects of Allopurinol on Beer-Induced Increases in Plasma Concentrations of Purine Bases and Uridine

We investigated the effects of allopurinol on beer-induced changes in the plasma concentration and urinary excretion of purine bases. Five healthy subjects underwent three studies: ingestion of beer after taking 300 mg allopurinol (combination study); ingestion of beer alone; ingestion of allopurinol alone. Increased plasma concentrations and urinary excretion of hypoxanthine were greater in the combination study than the beer alone study. However, increases in total plasma purine base concentrations were greater in the beer alone study, even though increases in plasma uridine concentrations did not differ. Beer-induced increases in plasma concentrations of purine bases appear partially offset by increased urinary excretion of hypoxanthine after allopurinol, which also controls increases in plasma uric acid levels caused by alcoholic beverage ingestion.

Effects of exercise and grape juice ingestion in combination on plasma concentrations of purine bases and uridine

Background Since grape juice contains considerable amounts of fructose, which may increase the plasma concentration of urate, the combination of exercise and grape juice may increase the plasma concentration of urate to a greater degree than grape juice or exercise alone. Methods We performed 3 experiments with 6 healthy male Japanese. The first was exercise alone (exercise alone experiment), the second was grape juice ingestion alone (grape juice alone experiment), and the third was a combination of exercise and grape juice ingestion (combination experiment). Results In the exercise alone experiment, the concentrations of purine bases and uridine in plasma, and lactate in blood, as well as the urinary excretion of oxypurines were increased, whereas the urinary excretion of uric acid and fractional excretion of purine bases were decreased. In the grape juice alone experiment, the concentrations of purine bases and uridine, as well as lactate in blood were increased, whereas the fractional excretion of uric acid was decreased. In the combination experiment, the concentrations of purine bases and uridine in plasma, and lactate in blood, as well as the urinary excretion of oxypurines were increased, whereas the urinary excretion of uric acid and fractional excretion of hypoxanthine, xanthine, and uric acid were decreased. The increase in plasma concentration of urate by the combination of exercise and grape juice was greater than that by each alone, though it was not significantly different from the sum of increases in those 2 experiments. Conclusion Increases in adenine nucleotide degradation and lactic acid production caused by both exercise and grape juice ingestion play an important role in the increase in plasma concentration of urate, while those in combination have an additive effect on that concentration.

原著3『食品におけるプリン体含有量の測定-和食の食材および健康食品について』

食品中のプリン体含量を測定した.今回対象とした食品は,和食で使用される野菜を中心とした食材で今までに測定されていない食品とおつまみ類,および健康食品である.プリン体含量は,四塩基合計値として,空豆,おくら,もやしは40mg/100g以下であった. 豆もやし, 貝割れ大根,おから,ブロッコリースプラウト,舞茸は50～130mg/100gとpurine-rich vegetablesに分類された.おつまみ類(生ハム,さきいか,アーモンド),調味料(唐揚げ粉),粉末スープ類(ポタージュ, コンソメ) には3 0～180mg/100gのプリン体が含有されていた.一方,健康食品(ケール,ローヤルゼリー, ビール酵母, クロレラ,DNA/RNA)は40～21500mg/100gと大量のプリン体を含有するものがあった.今回測定した食品では,健康食品に,非常に多くのプリン体を含むものがあり,推奨される1日のプリ体摂取量400mgの半量を占めることから,これらの健康食品の日常的な服用には注意が必要であると考えられた

The response of gastrointestinal microbiota to avilamycin, butyrate, and plant extracts in early-weaned pigs1,2

An experiment was designed to evaluate the effects of 3 different additives on the gastrointestinal microbiota of early-weaned pigs. Early-weaned (18 to 22 d; n = 32) pigs (6.0 +/- 0.10 kg of BW) from 8 litters were randomly distributed into 8 pens. Each pen was assigned 1 of 4 dietary treatments: a prestarter or control diet, the control diet with 0.04% avilamycin (AB), with 0.3% sodium butyrate, or with 0.03% plant extract mixture (XT; standardized mixture with 5% (wt/wt) carvacrol extracted from Origanum spp., 3% cinnamaldehyde extracted from Cinnamonum spp., and 2% capsicum oleoresin from Capsicum annum). At the end of the experimental period, 8 pigs per treatment were killed, and samples of their intestinal content were taken. The total bacterial load along the gastrointestinal tract (GIT; stomach, jejunum, cecum, and distal colon) and the lactobacilli and enterobacteria in the jejunum and cecum were measured by quantitative PCR. The total microbial counts along the GIT did not differ among the diets, but there was an increase in the lactobacilli:enterobacteria ratio in the cecum of the piglets on the XT diet (P = 0.003). Restriction fragment length polymorphism of the PCR-amplified V3, V4, and V5 regions of the 16S rDNA gene showed changes in the structure of the microbial community in the jejunum. Dendrograms grouped animals by diets; control with 0.3% sodium butyrate was the treatment that promoted the biggest changes in the microbial ecosystem, followed by AB and then XT. Biodiversity increased when using additives compared with the control diet (P = 0.002). Microbial metabolic activity along the hindgut was studied using the concentration of purine bases and carbohydrase activities. Different patterns for purine bases were observed between diets (diet x intestinal section, P = 0.01). The control diet reached a maximum purine base concentration at the end of the colon, whereas that of the AB diet was reached at the cecum. We could not detect any cellulase or xylanase activities in animals of this age. Appreciable amylase and amylopectinase activities were found, but they did not differ between diets. The results suggest that the effects of avilamycin, butyrate, or the plant extract would not be related to a reduction in the number of total bacteria inhabiting different sections of the GIT but rather to changes in the ecological structure and metabolic activity of the microbial community.

Effects of Allopurinol on Beer-induced Increases in Plasma Concentrations and Urinary Excretion of Purine Bases (Uric Acid, Hypoxanthine, and Xanthine)

To determine the effects of allopurinol on beer-induced increases in plasma and urinary excretion of purine bases (hypoxanthine, xanthine, and uric acid), we performed three experiments on five healthy study participants. In the first experiment (combination study), the participants ingested beer (10 ml/kg body weight) eleven hours after taking allopurinol (300 mg). In the second experiment (beer-only study), the same participants ingested beer (10 ml/kg body weight) alone, while in the third experiment (allopurinol-only study), they took allopurinol (300 mg) alone. There was a two-week interval between each of the studies. Beer-induced increases in plasma concentration and urinary excretion of hypoxanthine in the combination study were markedly higher than those in the beer-only study. On the other hand, the sum of increases in plasma concentrations of purine bases in the beer-only study was greater than in the combination study, whereas the increase in plasma uridine concentration in the combination study did not differ from the beer-only study. In addition, allopurinol administration inhibited the beer-induced increase in plasma concentration of uric acid. These results suggest that abrupt adenine nucleotide degradation may increase plasma concentration and urinary excretion of hypoxanthine under conditions of low xanthine dehydrogenase activity, which is mostly ascribable to allopurinol. Further, the difference in the sum of increases in plasma concentrations of purine bases between the combination study and beer-only study was largely ascribable to a greater increase in urinary excretion of hypoxanthine in the combination study. In addition, allopurinol intake seems to be effective in controlling the rapid increase in plasma uric acid caused by ingestion of alcoholic beverages.

『食品におけるプリン体含有量の測定-大豆加工食品を中心に多様化する各種食品について』

Concentrations of purine bases in soybean-derived food and in some Japanese food were quantitatively determined with the use of high-performance liquid chromatography (HPLC). After hydrolyzation with 70% perchloric acid, each of the bases, adenine, guanine, hypoxanthine and xanthine were determined. Total purine bodies were calculated by the sum of each of the purine bases. Concentrations of purine bodies were as follows: food derived from soybeans: 21.9 - 63.5 mg/100g or 100mL; purine-rich vegetables: 39.1 - 171.8 mg/100g; beers: 0.18 - 8.26mg/100mL; Japanese snacks: 14.1 - 159.3 mg/ 100g or 100mL. It is recommended that the amount of dietary purine bodies consumed by a gouty or hyperuricemic patient be limited to 400 mg per day. Although purine concentrations in the food reported here did not exceed 200 mg/100g, overeating should be avoided and good dietary habits are commendable.

Diet and procedures used to detach particle-associated microbes from ruminal digesta influence chemical composition of microbes and estimation of microbial growth in Rusitec fermenters.

Four different detachment methods were evaluated for their ability to remove particle-associated microorganisms (PAM) from ruminal digesta in semicontinuous fermenters fed two diets differing in their forage:concentrate ratio (80:20 [C20] and 20:80 [C80]). In the methylcellulose method, ruminal digesta was incubated at 38 degrees C for 15 min with saline solution containing 0.1% methylcellulose before being stored at 4 degrees C for 24 h. In the other procedures, samples were incubated with 0.1% methylcellulose before storage for 24 h at 4 degrees C in different solutions (pH = 2): 1) saline solution with 0.1% Tween 80; 2) saline solution with 0.1% Tween 80 and 1% tertiary butanol; and 3) saline solution with 0.1% Tween 80, 1% methanol, and 1% tertiary butanol. Common to all treatments was subsequent homogenization, followed by filtration and resuspension of the residue five times in the treatment solutions. Microbial removal was estimated indirectly by measuring removal of 15N. There were no differences (P > 0.05) among detachment procedures, neither in the detaching efficiency (mean values of 79.7 and 88.1% for C20 and C80 diets, respectively) nor in the total recovery of PAM (54.9 and 34.9% for C20 and C80, respectively). There were no differences (P > 0.05) among PAM pellets obtained by the different detachment procedures in their N content, purine bases (PB) concentration, or PB:N ratio. For the C80 diet, 15N enrichment was greater (P < 0.05) in PAM pellets obtained with methylcellulose than in those obtained by the other methods. However, there were no differences (P > 0.05) due to the detachment procedure in the values of daily microbial growth estimated using as reference the different PAM pellets. The PAM pellets for diet C20 presented greater (P < 0.01) N content and lower (P < 0.01) PB concentration than those for diet C80 (mean values of 74.3 vs 49.1 mg of N/g of dry matter, and 22.8 vs 26.0 micromol PB/mg of dry matter, respectively). Daily microbial growth was greater (P < 0.05) for the C80 diet than for the C20 diet (121 vs 114 mg of microbial N, respectively). Results suggest that the treatment of ruminal digesta with a saline solution with 0.1% methylcellulose at 38 degrees C for 15 min combined with homogenizing and chilling at 4 degrees C for 24 h removed a major proportion of PAM, although further research is needed to decrease microbial losses during the isolation process.

Consumption of Raw Potato Starch Increases Colon Length and Fecal Excretion of Purine Bases in Growing Pigs

Male growing pigs were fed a diet containing 250 g/kg of native corn starch (CS; 26% amylose, 74% amylopectin) or 250 g/kg of raw potato starch (RPS), as examples of digestible starch and resistant starch (Type II), respectively. Whole-tract digestibilities of organic matter, crude protein and starch were greater in pigs fed CS than in those fed RPS through at least d 23 of the study. However, the values progressively increased in the RPS-fed pigs up to d 38, at which time the groups did not differ in organic matter and starch digestibility. The digestive tract and colonic digesta were heavier and colon length longer in pigs fed the RPS diet. Digestibility of starch in the ileum on d 38 was significantly lower in RPS-fed pigs, but rose from ileum to rectum; most starch was extensively fermented in the cecum and proximal colon. Purine base (PB) and short-chain fatty acid (SCFA) concentrations in feces initially increased and then decreased beginning on d 4 for PB and on d 21 for SCFA. PB concentration in feces was greater in pigs fed RPS than in those fed CS. In the large bowel digesta, PB and SCFA concentrations increased from the ileum to the cecum and proximal colon and then fell in the distal colon. Pigs fed the RPS diet had a higher PB concentration in the middle colonic digesta and a greater SCFA concentration in the proximal colonic digesta than the CS-fed group. Adaptation of growing pigs to supplementary RPS required 5 wk, as reflected by whole-tract digestibility and PB and SCFA fecal excretion data.

Comparison of microbial markers (15N and purine bases) and bacterial isolates for the estimation of rumen microbial protein synthesis

AbstractThe first objective of this experiment was to investigate the effect of using different bacterial isolates on the estimation of microbial protein production in semi-continuous fermenters (RUSITEC) given four diets, and to test whether a ‘mixed’ bacterial pellet was representative of the whole bacterial population in the fermenters. A second objective was to compare two different microbial markers (nitrogen-fifteen (15N) and purine bases (PB)). Diets consisted of neutral-detergent fibre from grass hay (10 g/day) and sugar-beet pulp (2 g/day) and 280 mg/day of one of four N forms (isolated soya-bean protein, soya-bean peptides, amino acids blended to profile soya-bean protein and NH4Cl). Two 14-day incubation runs were carried out and in each run each of the four different diets were given to two vessels. On days 12 and 13, total digesta (effluent plus nylon bags residues) was collected for analyses of non-ammonia N,15N enrichment and PB concentration, and for isolation of total mixed bacterial pellets (TB). On the last day of each incubation run, the system was stopped for isolation of liquid- (LAB) and solid-associated (SAB) bacteria. Microbial N flow was estimated from the15N enrichment and PB concentration in both total digesta and in the three different bacterial pellets (TB, LAB, and SAB). For all diets, LAB presented a greater (P &lt; 005)15N enrichment and PB: N ratio than SAB, with TB having an intermediate value. For both markers, the use of LAB produced the lowest (P &lt; 005) estimates of microbial N flow and the use of SAB produced the greatest (P &lt; 005) estimates. The use of TB produced intermediate values with all diets, suggesting that TB consisted of SAB and LAB. For all bacterial pellets, PB produced greater (P 005) values of microbial N flow than15N. However, there was a positive relationship (r = 0·883; P 0001; no. = 15) between the values of microbial N flow determined with the two markers when TB were used as reference.

Influence of β-Mannase on Broiler Performance, Digestibility, and Intestinal Fermentation

Despite the increasing availability of feed enzymes and a growing interest in soybean meal (SBM) carbohydrates, there is little information on the potential beneficial effects of dietary supplements of enzymes to increase nutritive value. Here we hypothesize that a combination of mannase (polysaccharidase) and protease enzymes could reduce the anti-nutritive effect described in soybean β-manans and consequently increase the degradability of carbohydrates and proteins in diets based on SBM. We assessed the effect of adding mannase and protease to a corn-SBM diet fed to broiler chicks in batteries, particularly on productive performance and nutrient digestibility. Our results showed that neither BW gain nor feed intake was affected. No consistent effects were observed with the individual enzymes (mannase or protease) on the ileal or fecal digestibility of crude protein and crude fat. However, organic matter digestibility and apparent metabolizable energy of diets tended to increase with mannase supplementation. Addition of this enzyme to the diet also reduced ileal microbial flora (as measured by purine base concentration), which indicated that feed enzymes might contribute to the prevention of digestive diseases.