LACTOFERRIN SUPPLEMENTATION AND DIARRHEA

Abstract

The present invention relates in general to the field of intestinal health. In particular, the present inventors have found that a composition comprising lactoferrin can be used in the prevention, amelioration, or treatment of diarrhea. This finding is in particular important because long lasting diarrhea is seen as a common cause of mortality in infants and children.

Claims

1. A method for reducing the severity of diarrhea, the method comprising: administering a composition comprising lactoferrin to a subject in need of same.

2. The method in accordance with claim 1, wherein the diarrhea is a non-infectious diarrhea.

3. The method in accordance with claim 1, wherein the diarrhea is a weaning diarrhea.

4. The method in accordance with claim 1, wherein the lactoferrin is provided as a milk fraction.

5. The method in accordance with claim 1, wherein the subject is an infant or child.

6. The method in accordance with claim 1, wherein the composition comprises lactoferrin in an amount corresponding to about 5-500 mg Lactoferrin/kg body weight/day.

7. The method in accordance with claim 1, wherein the composition comprises lactoferrin in an amount corresponding to about 100-300 mg Lactoferrin/kg body weight/day.

8. The method in accordance with claim 1, wherein the composition comprises greater than 2 g/L to 3 g/L lactoferrin.

9. The method in accordance with claim 1, wherein the composition is administered at least daily.

10. The method in accordance with claim 1, wherein the subject is a human or animal.

11. The method in accordance with claim 1, wherein the subject is an infant who has experienced at least one episode of a non-infectious diarrhea.

12. The method in accordance with claim 1, wherein the subject is an infant who has experienced at least one episode of a weaning diarrhea.

13. The method in accordance with claim 1, wherein the subject is a mammal during the first 38 days after birth.

14. The method in accordance with claim 1, wherein the composition is selected from the group consisting of an infant feeding formula, a nutritional formula, a formula for feeding newborn animals, a nutritional supplement, a food additive, a foodstuff and a drink.

15. The method in accordance with claim 1, wherein the severity of diarrhea is based on fecal consistency.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0052] FIG. 1 shows the effect of lactoferrin supplementation on the non-infectious diarrhea ratio in formula fed piglets during the first 38 days after birth.

[0053] FIG. 2 shows the survival curve for the incidence of non-infectious diarrhea in piglets during the first 38 days after birth. Lines with different letters were significantly different (P<0.05, Kaplan-Meier survival curve) based on Log-rank pairwise test.

[0054] FIG. 3 shows the mean days of duration weaning (non-infectious) diarrhea in formula fed piglets during the first 38 days after birth. Values are meansSEM of 18 piglets. The columns with different letters were significantly different (P<0.05) based on Univariate ANOVA Bonferroni adjustment for multiple comparisons.

[0055] FIG. 4 shows the effect of lactoferrin supplementation on the severity of weaning (non-infectious) diarrhea in formula fed piglets during the first 38 days after birth (meansSEM).

[0056] FIG. 5 shows that lactoferrin supplementation significantly reduces the mean incidence of non-infectious diarrhea in formula fed piglets with dose response during the first 38 days of life P=0.017, General linear model (Univariate ANOVA) with Bonferroni's adjustment for the use of antibiotic as a covariance). Values are meansSEM of 18 replicate piglets. The columns with different letters were significantly different (P<0.05) based on Bonferroni adjustment.

EXAMPLES

Experimental Procedures

Animals

[0057] Seventy-two 3-day-old male domestic piglets (Sus scorfa LandraceLarge White F1) from 5 litters were purchased from a commercial piggery in Xiamen. The piglets were randomly allocated to 1 of 4 treatments according to weight and litter. All animals were housed in pairs in a temperature controlled environment with a 12-h light (08:00-20:00) and dark (20:00-08:00) cycle. The home pens contained a nest (a rubber tire covered with a clean towel), a heat lamp over the nest and an identical woody toy hang in the home pen. The maximum capacity of holding piglets at the behavior lab were 16, thus 5 trials (10-16 piglets/trial) were conducted to reach 16-18 piglets/group (Tab 1). The piglet behavior (including home pens) were monitored for 24-h with 7 camera surveillance systems. Two 3 day-old piglets were collected in each trial and euthanized as baseline control (n=10). The study protocol was approved by the Xiamen University Animal Ethics Committee.

TABLE-US-00001 TABLE 1 Grouping assignment Group Group 1 Group 2 Group 3 Group 4 Behavioral test Yes Yes Yes No Dosage Control Low High High Total 18 18 18 18 piglet/group

Lactoferrin Supplementation

[0058] Bovine milk lactoferrin was purchased from DMV international. Piglets were fed a standard sow milk replacer containing protein of soy/whey/casein (50:38:12) from 3 days old to 38 days old. The amount of lactoferrin in the final milk varied according to the groups: 0.05 g/L (group 1, the control group with no added lactoferrin), 0.5 g/L (group 2, sufficient dose), 1 g/L (group 3, high dose). All piglets in groups 1-3 were exposed to learning challenges. Group 4 was supplemented with lactoferrin at the same dose as group 3, 1 g/L, but was not exposed to learning challenges (as sham group).

[0059] These concentrations represented an approximate intake of lactoferrin in the control, sufficient dose and high dose group were 15, 145 and 285 mg/kg body weight/day, respectively. The pig milk replacers were formulated so that total protein intake remained the same irrespective of the amount of added lactoferrin. To maintain normal growth rates, the piglets received 285 mL milk/kg body wt/day in the first 2 wk of the study, and 230 mL/kg body weight/day in the remaining weeks. Feeding times were at 08:00, 13:00, 18:00, and 22:30, with an extra 50 mL milk/pig supplied at the last feeding. Body weight was weighted every morning before feeding using a digital scale (PRIS-Scale model: XK 3116). The milk intake, health status, stool frequency, stool score, medication of piglets were recorded daily. Diarrhea was defined as having three or more stools than usual, with very loose or mucous.

Evaluated Parameters

1. Incidence of Diarrhea

[0060] It is calculated as (number of piglets with diarrhea/total number of piglets in each group)100%

2. Duration of Diarrhea

[0061] It is defined as total length (days) of diarrhea within a group

3. Severity of Diarrhea

[0062] Severity of diarrhea was quantified using a faecal consistency score based on a scale from 0 to 3 (0=normal faeces, 1=soft faeces, 2=soft faeces with white or yellow mucus, 3=watery diarrhea)

4. Mean Incidence of Diarrhea

[0063] It is defined as the total number of diarrheas occurred in each piglet within a group.

Statistical Analysis

[0064] Differences in incidence of diarrhea (ratio number of piglet's diarrhea vs non-diarrhea) were compared using Chi-square Tests. The differences in the onset of diarrhea were compared using Kaplan-Meier survival analysis with Cox regression to examine potential covariates that may influence diarrhea. Comparisons between means of the average duration, severity, and average incidence of diarrhea in different groups were performed using the general linear model (Univariate ANOVA) with Bonferroni adjustment for multiple comparisons where appropriate. All statistical analyses were completed using SPSS for Windows 11 and 12 Inc, Chicago. A significance level of 0.05 was used.

Results

[0065] 1. Lactoferrin supplementation reduces the incidence of diarrhea in formula fed piglets during the first 38 days after birth FIG. 1 shows the effect of lactoferrin supplementation on the non-infectious diarrhea ratio in formula fed piglets during the first 38 days after birth. The diarrhea ratio was calculated as the percentage of piglets that had symptoms of diarrhea during the first 38 days after birth. The diarrhea ratio has a trend to decrease with the increase of lactoferrin supplementation concentration.

[0066] 2. Lactoferrin supplementation significantly delayed the onset of diarrhea in formula fed piglets during the first 38 days of life using Kaplan-Meier survival curve analysis (P<0.05, FIG. 2). Compared with control group, the onset of non-infectious diarrhea was delayed in formula fed piglets in response to lactoferrin in a dose-dependent manner (P=0.044, Log-rank test).

[0067] 3. Lactoferrin supplementation reduces the duration of weaning (non-infectious) diarrhea in formula fed piglets during the first 38 days after birth. The duration of diarrhea was calculated as the average days of diarrhea symptoms occurred in each group. Compared with control group (1.140.27 days), the duration of weaning (non-infectious) diarrhea decreased to 0.460.17 days with learning challenge and 0.640.23 day without learning challenge group when we administrated lactoferrin at dose level of 1 mg/L.

[0068] Also there was significant dose responses effect in reducing the duration of weaning (non-infectious) diarrhea after lactoferrin supplementation using General linear model (Univariate ANOVA) with Bonferroni's adjustment for multiple comparisons, antibiotic was used as the covariance (P=0.03, FIG. 3).

[0069] 4. Lactoferrin supplementation reduces the severity of weaning (non-infectious) diarrhea in formula fed piglets during the first 38 days after birth (FIG. 4). Severity of diarrhea was quantified using a faecal consistency score based on a scale from 0 to 3 (0=normal faeces, 1=soft faeces, 2=soft faeces with white or yellow mucus, 3=watery diarrhea and color 1=yellow, 2=yellow or white mucous stool, 3=watery stool).

[0070] 5. Lactoferrin supplementation reduces the mean incidence of weaning (non-infectious diarrhea) in formula fed piglets during the first 38 days after birth (FIG. 5). Compared with control group (1.060.21 day), supplemental lactoferrin reduces the incidence of weaning (non-infectious) diarrhea in formula fed piglets (0.560.25 day in 1.0 mg/L with learning group, and 0.560.18 day in 1.0 mg/L without learning group) during the first 38 days after birth, and the dose response was significant (P=0.017) based on the General linear model (Univariate ANOVA) with Bonferroni's adjustment for multiple comparisons, the use of antibiotic was taken as a covariance.

CONCLUSION

[0071] Dietary supplementation of lactoferrin not only decreased the incidence and severity of diarrhea, but also delayed the onset of diarrhea for newborn piglets. These findings show that dietary lactoferrin supplementation improves gut comfort and is beneficial for the gut maturation process for young mammals.