Current situation and perspectives of ready-to-eat food/meal suppliers.

Working Group 2 of the Healthy Diet Research Committee of International Life Sciences Institute Japan (WG2) assessed the concept and practice of healthy eating in the ready-to-eat food/meal industry in Japan. WG2 interviewed 14 arbitrarily selected member companies that included "health" or "nutrition" in their management policy, and sent a questionnaire to 338 member companies of the Japan Ready-Made Meal Association. Ready-to-eat food/meal suppliers mainly referred to Dietary Reference Intakes for Japanese, the Japanese Food Guide, and/or Healthy Japan 21 for their menu construction. They increased dietary fiber, variety, vegetables, whole-grain cereals, millet rice, and soy bean products; and reduced energy, carbohydrates, and salt in "healthy" food. They tended to avoid making direct appeals to health. Many companies reduced the salt content without drawing attention to the practice. They continually strive to improve flavor as the single most important factor for selling healthy food. The cycling of menus is used to increase diversity in food consumption. These industries require both academia and the government to define priorities for increasing and decreasing particular nutrients as the main targets and to establish the maximum time for balancing each nutrient.

Comparison between the effects of ultrasound and gamma-rays on the inactivation of Saccharomyces cerevisiae: analyses of cell membrane permeability and DNA or RNA synthesis by flow cytometry.

The effects of 200 kHz ultrasonic irradiation on DNA or RNA formation and membrane permeability of yeast cells were investigated by flow cytometry and compared with those of (60)Co gamma-ray radiation. Colony counting analyses were also performed for comparison. It was observed that the colony-forming activity of yeast cells was not affected by small doses of ultrasonic irradiation, but was closely related to the amounts of sonolytically formed hydrogen peroxide at concentrations of more than 80 microM. On the other hand, gamma-rays directly retarded colony-forming ability in addition to the effects of radiolytically formed hydrogen peroxide. The results obtained by flow cytometry also indicated that the amounts of DNA or RNA formed decreased with an increase in ultrasonic irradiation time without any threshold. These results indicated that flow cytometry can show early growth activities, but that colony counting analyses are insufficient to evaluate continuous and quantitative changes in these activities. In addition, by analyzing the amounts of DNA or RNA formed in the presence of the same amount of hydrogen peroxide, it was found that DNA or RNA formation behavior in the presence of hydrogen peroxide with no irradiation was similar to that following ultrasonic irradiation. These results suggested that similar chemical effects due to the formation of hydrogen peroxide were produced during ultrasonic irradiation. In addition, physical effects of ultrasound, such as shock wave, hardly contributed to cell inactivation and cell membrane damage, because relatively high frequency ultrasound was used here. In the case of gamma-ray radiation, direct physical effects on the cells were clearly observed.

Pitfalls in short-chain fatty acid research: A methodological review.

This methodological review suggests what to do and what not to do in short-chain fatty acid (SCFA) research for researchers, supervisors, scientific reviewers, and regulatory officers. High viscosity of gut contents, existence of bacterial biofilm and of mucus layer at the mucosal surface, and rapid absorption of SCFAs make it difficult to know their concentrations at the very surface of the mucosa. As lumen or fecal concentration of SCFAs does not reflect their rate of production, these parameters should not be used as measures of SCFA production or absorption. Effects of SCFAs can vary and even become opposite at different dose, time of/after exposure or time of the day. Thus, results without dose-response, time-course, and diurnal variance experiments can be seriously misleading. It is also to note that too much emphasis on n-butyrate should be avoided.

Estimation of daily selenium intake by 3- to 5-year-old Japanese children based on selenium excretion in 24-h urine samples.

To evaluate the daily Se intake of 3- to 5-year-old Japanese children, we used seventy-two urine samples collected from fifty-three children (twenty-seven male and twenty-six female) from two cities in Miyagi prefecture, Japan. For measuring low Se concentrations with high precision, accuracy and rapidity in the 24-h urine samples, we developed an instrumental neutron activation analysis (INAA) method, that is without any chemical separation, using the short-lived 77mSe (t1/2 = 17·4 s) nuclide. The estimated Se intake of the fifty-three children was 51·5 (sd 30·2) µg/d (geometric mean: 42·7 µg/d). Ten subjects (three male and seven female), successfully provided 24-h urine samples over two or three consecutive days; their Se intake was 37·4 (sd 5·9) µg/d. Based on the logarithmically transformed data of these ten subjects, the ratio of intra-/inter-individual variances of usual Se intake was 16·7 (28·0/1·7) and geometric mean was 27·7 µg/d. The 5th to 95th percentile of usual Se intake of these ten subjects was 17·5 to 40·4 µg/d, which ranged between the recommended dietary allowance and tolerable upper intake level of Se by the Dietary Reference Intakes for Japanese (2015).

Insoluble dietary fiber of wheat bran increased viscosity of pig whole cecal contents in vitro.

Viscosity of whole pig cecal contents with (2 g/200 mL) or without insoluble dietary fibers prepared from wheat bran of two particle sizes (WB fine 0.22+/-0.22 mm(2), n=1,723; WB coarse 0.51+/-1.12 mm(2), n=1,457, mean+/-SD) was measured using a rotary viscometer equipped with a vane spindle at the shear rates of 0.05, 0.16, 0.47, 0.78 and 1.56 (s(-1)) and at 0 and 2 h. Such measurements were repeated twice each for five different donor pig groups. We were able to measure the coefficient of viscosity of such samples over a wide range of shear rate. Coefficient of viscosity depended on shear rate. Addition of insoluble dietary fiber increased the coefficient of viscosity. Insoluble dietary fiber from WB coarse had a significantly stronger effect than that from WB fine. The above results demonstrated that insoluble dietary fibers of wheat bran increase the viscosity of gut contents, and thereby potentially retard digestion and absorption.

Inhibitory effect of succinic acid on epithelial cell proliferation of colonic mucosa in rats.

Microbial breakdown of carbohydrates in the large intestine mainly produces short-chain fatty acids (SCFA). SCFA stimulate epithelial cell proliferation of the digestive tract in vivo. Succinic acid sometimes accumulates in the colonic lumen. However, the effect of succinic acid on colonic epithelial cell proliferation is unknown. Thus, we planned to clarify the influence of succinic acid on colonic epithelial cell proliferation in vivo. We continuously administered infusate with or without succinic acid (100 mM) into the distal colon of rats for 6 d and measured accumulated mitosis per crypt of distal colon of these rats. Succinic acid infused into rat colons significantly inhibited colonic cell proliferation and reduced crypt size. These results clearly indicated the inhibitory effects of succinic acid on colonic epithelial cell proliferation in vivo.

Influence of temperature on short-chain fatty acid production by pig cecal bacteria in vitro.

We studied the effect of incubation temperature on the production of shortchain fatty acids (SCFA) by pig cecal bacteria in vitro in order to assess short-term influences of body temperature on bacterial metabolism in the large intestine. We employed a 200 mL scale continuous culture system using cecal bacteria from commercially slaughtered pigs as innoculum. The culture was maintained at 30, 37, 40 or 42 degrees C and continuously diluted by continuous feeding of bicarbonate buffer (pH 7.4) added with lactose (10 g/L) and by simultaneous continuous efflux both at 4.17 mL/h. We monitored SCFA concentration of the culture for 12 h, which represents their production rate. Concentrations of SCFA increased during the first several hours and plateaued at around 11 h of incubation. Incubation temperature significantly affected mean concentrations from 1 to 12 h of acetic (40 degrees C>42 degrees C= 37 degrees C>30 degrees C), propionic (40 degrees C>42 degrees C=30 degrees C), n-butyric (42 degrees C>37 degrees C>30 degrees C, 40 degrees C>30 degrees C) and n-valeric (42 degrees C=40 degrees C>37 degrees C>30 degrees C) acids, and total SCFA (40 degrees C>42 degrees C= 37 degrees C> 30 degrees C) (p<0.05). These results indicate that both hyperthermia and hypothermia depress the microbial breakdown of carbohydrates.

Dose-dependent stimulatory and inhibitory effects of luminal and serosal n-butyric acid on epithelial cell proliferation of pig distal colonic mucosa.

Large bowel bacteria convert various carbohydrates into short-chain fatty acids (SCFA). SCFA stimulate epithelial cell proliferation of the large intestine in vivo and inhibit that of various cells in vitro. Supposing that too high concentration of SCFA on the serosal side is responsible for their inhibitory effect in vitro, we studied effects of luminal and serosal n-butyric acid (0, 0.1, 1, or 10 mmol/L, adjusted to neutral pH) on the epithelial cell proliferation rate of pig colonic mucosa in organ culture taking crypt cell production rate (CCPR) as the measure of proliferative activity. With 0 or 0.1 mmol/L n-butyric acid on the serosal side, luminal n-butyric acid increased CCPR at 1.0 mmol/L, and decreased CCPR at 10 mmol/L when compared to the luminal 0 mmol/L control. With 1.0 or 10 mmol/L serosal n-butyric acid, luminal n-butyric acid depressed CCPR dose-dependently. The above results indicated that n-butyric acid stimulated colonic epithelial cell proliferation at low concentration and inhibit it at high concentration with interaction effect to enhance the inhibitory action. The stimulatory effect of a low dose of serosal n-butyric acid may be responsible for the distant trophic effect of SCFA.

Histochemical structure of the mucus gel layer coating the fecal surface of rodents, rabbits and humans.

The aim of this study was to investigate whether a mucus layer covers the surface of various animal feces and, if so, to show the structure and mucin composition of this layer. The freshly excreted feces of mice, rats, guinea pigs, rabbits and humans were fixed with Carnoy solution. Cross-sections approximately 5 microm thick were stained with alcian blue (AB) or the periodic acid Schiff (PAS) reaction. We measured the thickness of the mucus layer on the fecal surface of these sections. The fecal surface was covered with a continuous mucus layer in all specimens. The mucus layers of mice, rats and humans consisted of the alternate stratification of AB-positive and PAS-positive mucin layers. In contrast, the mucus layer consisted of an inner PAS-positive neutral mucin layer and an outer AB-positive acidic mucin layer in guinea pigs and rabbits. The average thicknesses of the mucus layers upon the fecal surfaces for mice, rats, guinea pigs, rabbits and humans were 19 +/- 12, 22 +/- 14, 15 +/- 4.6, 19 +/- 14 and 17 +/- 11 microm, respectively. These results demonstrated that the fecal surfaces are covered with continuous mucus layers in rodents, rabbits and humans, with substructures varying among species.

Viscous properties of pig cecal contents and the contribution of solid particles to viscosity.

OBJECTIVE: It is believed that solid particles are not responsible for digesta viscosity. However, solid particles are responsible for the viscosity of whole blood and fiber suspension such as wood pulp. Therefore, we measured viscosity of gut contents with and without solid particles. We also determined the contribution of solid particles to viscoelastic characters of gut contents. METHODS: We measured the coefficient of viscosity, shear rate, and shear stress of pig cecal contents by using a tube-flow viscometer before and after removal of particles. RESULTS: The coefficient of viscosity of intact pig cecal contents was negatively correlated with shear rate, a typical character of non-Newtonian fluids. The removal of solid particles from the contents resulted in a Newtonian fluid with very low viscosity. CONCLUSION: Solid particles were mainly responsible for the viscoelastic character of pig cecal contents. Accordingly, the contribution of indigestible solid food components, sloughed epithelial cells, and bacterial cell bodies to the viscosity of gut contents should be re-evaluated.

Daily salt intake of healthy Japanese infants of 3-5 years based on sodium excretion in 24-hour urine.

We measured concentrations of sodium and creatinine in previously validated overnight urine and 24-h urine of 79 healthy infants of 3 to 5 y of age in two cities in Miyagi Prefecture, Japan. Thereafter, we estimated daily salt intake from daily urinary sodium excretion and daily urinary volume. Corrections for potential sweat loss of sodium chloride were conducted based on urinary volume and urinary creatinine concentration. We also measured urinary volume and urinary sodium concentration in infants who failed to provide complete urine samples. The estimated salt intake did not differ between boys and girls, but varied between the two cities. The estimated salt intake per unit of body weight differed neither between genders nor between cities. The grand means for estimated salt intake and that per unit of body weight were 5.5 g/d (SD 1.8) and 0.32 g/kg/d (SD 0.10), which supported previous results both in Japan and abroad. It was also possible to roughly estimate salt intake from sodium concentration in overnight urine or incomplete 24-h urine, and published urine volume.

Is glycemic index of food a feasible predictor of appetite, hunger, and satiety?

This review assesses the feasibility of using glycemic index (GI) as a predictor of appetite, hunger and satiety by surveying published human intervention studies. We also discuss the relationship between GI and two appetite/satiety control hormones, leptin and ghrelin. Ingestion of high-GI food increased hunger and lowered satiety in short-term human intervention studies. This effect may be attributed to the rapid decline in blood glucose level following a hyperinsulinemic response caused by a sharp and transient increase in blood glucose level that occurs after the ingestion of high-GI food, which is defined as the glucostatic theory. However, appetite, hunger and satiety after the ingestion of foods with varying GI were inconsistent among long-term human intervention studies. From the few relevant long-term studies available, we selected two recent well-designed examples for analysis, but they failed to elicit clear differences in glycemic and insulinemic responses between high- and low-GI meals (consisting of a combination of different foods or key carbohydrate-rich foods incorporated into habitual diets). One of the reasons that these studies could not predict glycemic response to mixed meals is presumably that the GI of each particular food was not reflected in that of the mixed meals as a whole. Thus, it is difficult to conclude that the GI values of foods or mixed meals are a valid long-term predictor for appetite, hunger and satiety. Both insulin and insulin-mediated glucose uptake and metabolism in adipose tissue affect blood leptin concentration and its diurnal pattern. Circulating ghrelin level is suppressed by carbohydrate-rich meals, presumably via glycemia and insulinemia. Accordingly, low-GI foods may not necessarily increase satiety or suppress appetite and/or hunger because of the lack of insulin-mediated leptin stimulation and ghrelin suppression. However, insulin-mediated leptin stimulation and ghrelin suppression per se is not consistent among studies; thus we were not able to identify a clear relationship among GI, satietogenic leptin, and appetitic ghrelin.

Large particles increase viscosity and yield stress of pig cecal contents without changing basic viscoelastic properties.

The viscosity of gut contents should influence digestion and absorption. Earlier investigators measured the viscosity of intestinal contents after the removal of solid particles. However, we previously found that removal of solid particles from pig cecal contents dramatically lowered the viscosity of the contents. Accordingly, we examined the contribution of large solid particles to viscoelastic parameters of gut contents in the present study. We removed large particles from pig cecal contents by filtration through surgical gauze. Then, we reconstructed the cecal contents by returning all, one half or none of the original amount of the large particles to the filtrate. We measured the viscosity, shear stress and shear rate of these reconstructed cecal contents using a tube-flow viscometer. The coefficient of viscosity was larger when the large-particle content was higher (P < 0.01). Cecal contents behaved as a non-Newtonian fluid and showed an apparent Bingham plastic nature irrespective of large-particle content. We calculated the yield stress of these fluids assuming that the fluids behave as Bingham plastic. The yield stress of the cecal contents was greater (P < 0.05) when the large-particle content was higher. The above results indicated that large particles elevated the viscosity and yield stress of gut contents without changing their basic viscoelastic character. Integrating the present and our previous results, we conclude that it is likely that finer particles such as bacteria should provide non-Newtonian and apparent Bingham plastic characteristics to pig cecal contents.

Production of short-chain fatty acids and gas from various oligosaccharides by gut microbes of carp (Cyprinus carpio L.) in micro-scale batch culture.

We studied the metabolism of various oligosaccharides by carp (Cyprinus carpio) hindgut microbes by measuring gas productivity and organic acid production in gut contents using a 50-microl-scale batch culture system. Carp hindgut contents were incubated with 500 microg each of raffinose, lactosucrose, kestose, lactulose, gentiobiose, 4'-galactosyllactose and 6'-galactosyllactose and soybean-, xylo-, and isomalto-oligosaccharides or none (blank culture) at 25 degrees C for 6 h. The time-course of gas release from the culture (Y microl/culture) was expressed as an exponential function of incubation time (t) [Y=A+Bx(1-e(-kt))]; A, B and k are constants). Potential production of gas (A+B) from soybean-oligosaccharide and raffinose was larger than for the other saccharides except for kestose, and blank culture. The rate constant of gas (k) for lactosucrose was larger than that for isomalto- and xylo-oligosaccharide, lactulose, kestose or blank culture. Net production of total SCFA (sum of acetic, propionic and n-butyric acid weights) from cultures with soybean- and isomalto-oligosaccharides, raffinose, gentiobiose and lactosucrose was greater than that from blank culture. These results suggested that soybean-oligosaccharide and raffinose were potentially highly fermentable oligosaccharides for carp hindgut microbes. Chemical structures of oligosaccharides seem to play an important role in the fermentability. It is also likely that oligosaccharide utilization differs between mammals and teleosts.

Glucose but Not Polypeptone Reduces the Effect of Probiotic Preparations to Stimulate Carbohydrate Breakdown and Reduce Net Ammonia Production by Pig Cecal Bacteria In Vitro.

Probiotic preparations are used to prevent or treat diarrhea. Probiotic preparations increase the in vitro breakdown of carbohydrates and decrease that of protein by mixed cecal bacteria in the absence of readily fermentable materials. Diarrhea can increase the influx of readily digestible materials into the large intestine. Therefore, we compared production of organic acids and ammonia in batch cultures using pig cecal contents with or without probiotic preparations (Clostridium butyricum, Lactobacillus casei plus Bifidobacterium breve, or L. casei plus Enterococcus faecalis) and with or without readily fermentable substrates (glucose or polypeptone). Effects of additional substrates appeared earlier than those of probiotic preparations. Both probiotic preparations and glucose increased the production of most short-chain fatty acids and lactic acid but decreased those of ammonia and isovaleric acid. Effects of probiotic preparations and glucose were not additive. Effects of different preparations were alike. Polypeptone increased the production of ammonia and isovaleric acid independent of probiotic preparations. Effects of probiotics and glucose may be to provide either energy or carbon to the microbial ecosystem. Effects of glucose were probably mediated through their direct utilization by indigenous bacteria.

Gastric or rectal instillation of short-chain fatty acids stimulates epithelial cell proliferation of small and large intestine in rats.

Short-chain fatty acids stimulate gut epithelial cell proliferation in vivo, although the difference between oral and rectal routes is unknown. Accordingly, we examined the effect of oral or rectal administration of these acids. We instilled a mixture of acetic acid, propionic acid, and n-butyric acid (150, 60, and 60 mmol/liter, respectively; pH 6.5) or saline (270 mM, pH 6.5) into the stomach (2 ml) or rectum (1 ml) three times daily for five days in rats fed an elemental diet. We measured crypt cell production rate of the jejunum, ileum, and distal colon of these rats. The crypt cell production rate of these segments was higher in rats with gastric or rectal instillation of short-chain fatty acids than in saline controls. The rectal route was slightly more effective than the gastric route. The above results indicated that the instillation of short-chain fatty acids orally or rectally stimulated gut epithelial cell proliferation.

Influences of probiotic bacteria on organic acid production by pig caecal bacteria in vitro.

The mechanism of action of probiotics is largely unknown. A potential mechanism should be to increase the production of short-chain fatty acids (SCFA), known modulators of gut functions, by the bacterial ecosystem in the large intestine. The present paper reviews our recent studies in which the capacity of probiotic bacteria to increase the production of SCFA by pig caecal bacteria was investigated using batch-culture and continuous-culture techniques. All four commercial probiotic preparations and three strains of probiotic bacteria dose-dependently accelerated the net production of SCFA, succinic acid and lactic acid without changing the acid profile, and slowed the net production of NH4. Effects on organic acid production did not vary among different probiotic species. Neither probiotic preparations nor probiotic bacteria affected the organic acid production from glucose, gastric mucin, starch or lactose, or organic acids produced:added saccharide. Glucose abolished these effects of probiotic preparations. However, the capacity of probiotics to increase SCFA production was not modified by gastric mucin, starch or lactose. These results indicate that probiotic bacteria increase SCFA production by accelerating the breakdown of carbohydrates that are resistant to indigenous bacteria, and suggest that the concept of prebiotics in terms of SCFA production as a measure of probiotic function is arguable.

Immature granulocyte count after liver transplantation.

We evaluated the significance of immature granulocyte (IG) count during the clinical course after liver transplantation. We counted IG using the flow cytometric method with CD16, CD11b, and CD45 antibodies. Samples were obtained from 31 patients in the Department of Transplantation and Immunology, and we determined (i) the distribution of IG peak value, (ii) the distribution of IG peak time-points, (iii) the clinical background of patients with high IG, and (iv) the clinical course of high IG cases. We observed the appearance of IG (100/microl or higher) in the majority of the patients (23 out of 31 patients; 74.2%). The IG peak was detected on the 19th day after transplantation. We observed serious complications, such as melena, rejection, or severe infection, in high IG (500/microl or higher) cases. We observed instances of inflammation with low C-reactive protein (CRP) value in the presence of IG. We believe that IG is a useful marker to monitor inflammation.