Composition for use in the therapy of lactose intolerance or conditions arising from lactase deficiency

Abstract

The invention provides a composition for use in therapy of lactose intolerance or conditions arising from lactase deficiency, wherein the composition is a non-dairy solid dosage form comprising: (a) a lactase; and (b) one or more lactase-producing and/or lactase containing microorganisms selected from Lactobacillus delbrueckii ssp bulgaricus and Streptococcus thermophilus; wherein the lactase (a) is other than a lactase derived from the said one or more microorganisms (b); and wherein the one or more lactase-producing and/or lactase-containing microorganisms (b) are in dried form. Also provided are uncoated capsule and tablet compositions containing the lactase and the lactase-producing and/or lactase-containing microorganisms.

Claims

1. A solid dosage form which is suitable for oral administration and is selected from capsules, tablets, dragees, powders, pellets and granules, wherein the solid dosage form is not coated by an enteric coating and contains a lactase (a) derived from Aspergillus oryzae which is stable and active in a pH range of 3 to 6; the term stable meaning that the lactase (a) still has at least 30% of its activity after one hour at 37° C. at a pH in the range from 3 to 6, the lactase being present in an amount of from 2000 to 50,000 FCC units; and contains as lactase-producing or lactase-containing microorganisms, both Lactobacillus delbrueckii ssp bulgaricus and Streptococcus thermophilus, the amounts of Lactobacillus delbrueckii ssp bulgaricus and Streptococcus thermophilus being between 100 million and 200 billion colony forming units; wherein the lactase (a) is other than a lactase derived from the said microorganisms; and wherein the microorganisms are in dried form; and provided that the solid dosage form is other than a solid dosage form containing in combination two or more added herbal extracts together with exogenous lipase and exogenous protease.

2. A solid dosage form according to claim 1 wherein the lactase (a) exhibits peak activity at a pH in the range from 3 to 6.

3. A solid dosage form according to claim 1 wherein the lactase (a) is derived from a microorganism, and the solid dosage form is substantially free of the microorganism from which the lactase is derived.

4. A solid dosage form according to claim 1 wherein the solid dosage form is selected from capsules, tablets and dragees.

5. A solid dosage form according to claim 1 wherein the solid dosage form is in unit dosage form.

6. A solid dosage form according to claim 5 wherein the unit dosage form is a capsule.

7. A solid dosage form according to claim 1 which contains between 2000 and 12000 FCC enzyme units of the lactase (a).

8. A solid dosage form according to claim 1 wherein the composition contains from 100 million to 10 billion colony forming units of Lactobacillus delbrueckii ssp bulgaricus and from 100 million to 10 billion colony forming units of Streptococcus thermophilus.

9. A solid dosage form according to claim 1 which is substantially free of lactose.

10. A solid dosage form according to claim 1 wherein the Lactobacillus delbrueckii ssp bulgaricus is Lactobacillus delbrueckii ssp bulgaricus LB-VC 18 (deposition number: DSM 23320).

11. A solid dosage form according to claim 1 wherein the Streptococcus thermophilus is Streptococcus thermophilus ST-VC18 (deposition number: DSM 23319).

12. A solid dosage form according to claim 1 which contains between 100 million and 10 billion colony forming units of Lactobacillus delbrueckii ssp bulgaricus.

13. A solid dosage form according to claim 1 which contains between 100 million and 10 billion colony forming units of Streptococcus thermophilus.

14. A method for the therapy of lactose intolerance or conditions arising from lactase deficiency, which method comprises administering to a subject in need thereof an effective amount of a composition which is a solid dosage form according to claim 1.

15. A method according to claim 14 wherein the composition is in unit dosage form.

16. A method according to claim 15 wherein the unit dosage form is a capsule.

17. A method according to claim 14 wherein the composition contains between 2000 and 12000 FCC enzyme units of the lactase (a).

18. A method according to claim 14 wherein the effective amount of the composition contains from 100 million to 10 billion colony forming units of Lactobacillus delbrueckii ssp bulgaricus and from 100 million to 10 billion colony forming units of Streptococcus thermophilus.

19. A method according to claim 14 wherein the composition is substantially free of lactose.

20. A method according to claim 14 wherein the Lactobacillus delbrueckii ssp bulgaricus is Lactobacillus delbrueckii ssp bulgaricus LB-VC18 (deposition number: DSM 23320).

21. A method according to claim 14 wherein the Steptococcus thermophilus is Streptococcus thermophilus ST-VC18 (deposition number: DSM 23319).

22. A method according to claim 14 wherein the composition is other than a composition containing in combination two or more added herbal extracts together with exogenous lipase and exogenous protease.

Description

EXAMPLES

(1) The invention will now be illustrated, but not limited, by reference to the examples set out below.

Example 1

Capsule Compositions

(2) The following capsule compositions may be prepared by mixing lactase from Aspergillus oryzae, and lyophilised Lactobacillus bulgaricus and Streptococcus thermophilus.

(3) Capsule Composition A—Size 1 Capsule

(4) A size 1 gelatine capsule (volume 0.5 ml) obtainable from Capsugel BVBA, Bornem, Belgium is filled with 70 mg lactase (activity of the lactase is 50,000 FCC/g), 40 mg of lyophilised Streptococcus thermophilus (30 billion cfu/g), 30 mg of lyophilised Lactobacillus bulgaricus (50 billion cfu/g) and 200 mg dicalcium phosphate per capsule.

(5) A preferred strain of Streptococcus thermophilus is Streptococcus thermophilus ST-VC18 (deposition number: DSM 23319) and a preferred strain of Lactobacillus bulgaricus is Lactobacillus delbrueckii LB-VC18 (deposition number: DSM 23320).

(6) Capsule Composition A—Size 1 Capsule

(7) A size 3 gelatine capsule (volume 0.3 ml) obtainable from Capsugel BVBA, Bornem, Belgium is filled with 70 mg lactase (activity of the lactase 50,000 FCC/g), 30 mg Streptococcus thermophilus (30 billion cfu/g), 20 mg Lactobacillus bulgaricus (50 billion cfu/g) and 100 mg dicalcium phosphate per capsule.

(8) Capsule Composition C—Size 0 Capsule

(9) A size 2 gelatine capsule (volume 0.68 ml) obtainable from Capsugel BVBA, Bornem, Belgium is filled with 150 mg lactase (activity of the lactase 50,000 FCC/g), 100 mg Streptococcus thermophilus (30 billion cfu/g), 50 mg Lactobacillus bulgaricus (50 billion cfu/g) and 250 mg dicalcium phosphate.

(10) Capsule Composition D—Size 2 Capsule

(11) A size 3 gelatine capsule (volume 0.37 ml) obtainable from Capsugel BVBA, Bornem, Belgium is filled with 100 mg lactase (activity of the lactase 100,000 FCC/g), 50 mg Streptococcus thermophilus (30 billion cfu/g), 30 mg Lactobacillus bulgaricus (50 billion cfu/g) and 100 mg dicalcium phosphate.

Example 2

An Investigation of the Effect of a Combination of Lactase and Lactic Acid Bacteria on Lactose Digestion in Lactose Malabsorbers

(12) An investigation was carried out to compare the effects of lactase, lyophilized lactic acid bacteria and a combination of lactase and lactic acid bacteria on lactose digestion in lactose malabsorbers.

(13) The investigation consisted of a placebo-controlled, randomized, double-blind cross-over study involving twenty four lactose malabsorbing human subjects. Thirty subjects were initially recruited to the study but six were found not to be lactose malabsorbers and the results obtained from these six subjects were therefore excluded from the analysis of the results.

(14) The study comprising an entry examination and five experimental days, separated by 4 wash-out periods of two weeks each. During the study, the participants were asked not to change their dietary habits and to abstain from taking vitamins, minerals and other supplements. At the beginning of each experimental day, after an overnight fast, a first breath sample was taken from each subject. Thereafter each subject ingested one of five test preparations in randomised order.

(15) The test preparations were provided in capsules of identical appearance, of which one per experimental meal was administered to the study participants together with 150 ml milk to which 5 g of lactose was added, resulting in a lactose-fortified milk containing approximately 12.5 g of lactose. The compositions of the five test preparations were as follows:

(16) Preparation (a): Capsule containing 1×10.sup.9 cfu Streptococcus thermophilus plus 1×10.sup.9 cfu Lactobacillus delbrückii ssp. bulgaricus.

(17) Preparation (b): containing 3300 FCC acid lactase from Aspergillus oryzae

(18) Preparation (c): Capsule containing 9000 FCC acid lactase from Aspergillus oryzae

(19) Preparation (d): Capsule containing a combination of 3300 FCC acid lactase plus 1×10.sup.9 cfu Streptococcus thermophilus and 1×10.sup.9 cfu Lactobacillus delbrückii ssp. bulgaricus (test preparation a))

(20) Preparation (e): Di-Calcium-Phosphate (placebo).

(21) The lactic acid bacteria in test preparations (a) and (d) were the strains ST-VC 18 and LB-VC18 described in Example 1 and in the description on page 24 above. Test preparation (b) was Lactrase 3300 (Pro Natura Gesellschaft für gesunde Ernährung mbH, Bad Vilbel, Germany), test preparation (c) was Lactrase Plus 9000 (Pro Natura Gesellschaft für gesunde Ernährung mbH, Bad Vilbel, Germany) and test preparation (d) was Lactrase 3300 (Pro Natura) to which the lactic acid bacteria were added.

(22) The randomization of study participants as well as packaging and coding of the test preparations was performed by subjects who were otherwise not involved in the study.

(23) The lactose dose given (12.5 g) was less than for a classic lactose malabsorption test (50 g) but is closer to the lactose-content of normally ingested foodstuffs. In the classic lactose malabsorption test (carried out with 50 g of lactose) H.sub.2 peak concentrations of 20 ppm and more are considered to be a proof of lactose malabsorption. However, because of the smaller lactose doses given in the present study, a lower threshold was used to determine whether a participant was a lactose malabsorber. Therefore, participants with H.sub.2 peak concentrations of 13 ppm and more with placebo were considered lactose malabsorbers and the results obtained from these participants were included in the study.

(24) The effect of the test preparations on lactose digestion was measured using H.sub.2-breath tests which were carried out on a MicroLyzer Model 12i, QuinTron Instruments, Milwaukee, Wis. 53215, USA. Hydrogen gas is released in approximately 90% of the population when carbohydrates are fermented by bacteria in the colon. Therefore, the amount of hydrogen in exhaled breath provides a good indicator of the amount of lactose being fermented in the colon and also constitutes a good indicator of lactase deficiency in the small intestine.

(25) Breath samples were taken for 3 hours. The H.sub.2 peak concentration was chosen as a parameter for lactose digestion. The H.sub.2 peak concentrations for each of the subjects and each of the five preparations are shown in the table below.

(26) TABLE-US-00001 H.sub.2 Max values Subject Preparation Preparation Preparation Preparation Preparation ID (e) (a) (b) (c) (d) 3501 27 48 2 16 7 3503 15 12 4 2 18 3504 37 36 33 46 9 3505 27 10 24 16 15 3506 27 14 29 35 18 3507 48 44 25 14 7 3508 18 38 24 23 15 3509 22 33 45 27 20 3510 33 33 28 33 28 3511 29 29 12 45 18 3513 13 10 6 4 17 3514 17 9 8 10 4 3516 15 21 7 10 13 3517 34 19 10 3 10 3518 15 5 16 22 11 3520 15 7 14 10 6 3521 27 12 31 5 9 3522 13 5 8 2 16 3524 21 16 5 3 5 3526 26 18 16 16 19 3527 19 17 18 16 12 3528 22 15 18 12 13 3529 45 45 61 24 15 3530 53 33 22 39 43

(27) Statistical analysis of the results of the study showed that there was a significant global effect between the diets with respect to the H.sub.2-max values obtained (Friedmann-Test, p=0.001). Multiple comparison (Wilcoxon) showed significant differences with respect to H.sub.2-max values between preparation (b), which contains 3300 FCC units of lactase, and the placebo preparation (e), between (c), which contains 9000 FCC units of lactase, and the placebo preparation (e) and between the combination preparation (d), which contains 3300 FCC of lactase plus 2 billion cfu lactic acid bacteria, and the placebo preparation (e). Moreover the mean variation (dispersion) of the H.sub.2-max values, i.e. the 25/75%-interquartiles area for preparation (d) was only half as big as the mean variations of the other test preparations (i.e. preparations (a), (b) and (c)).

(28) The results demonstrated that all four preparations containing active ingredient(s); i.e. preparation (a) (bacteria alone), preparation (b) (3300 FCC), preparation (c) (9000 FCC) and preparation (d) (the combination preparation) reduced the H.sub.2-breath concentrations in at least some subjects but only the 3300 FCC preparation (preparation (b)), the 9000 FCC preparation (preparation (c)) and the combination preparation (preparation (d)) showed a significant effect. The combination preparation (d) was even more effective than the 9000 FCC-containing preparation (c) and also showed a significantly lower mean variation in the H.sub.2-exhalation-values. An analysis of the data showed that fewer participants who received the combination preparation (d) enjoyed little or no beneficial effect. The advantage of the combination preparation (d) was therefore not only its stronger contribution to lactose digestion but also that it was effective in a larger proportion of the study participants. It may therefore be concluded that more lactose malabsorbers would benefit from the lactose digestive effect of combination preparation (d) than would benefit from the individual lactic acid bacteria preparation (a) and the lactase preparations (b) and (c).

Example 3

An Investigation of Lactose Turnover by Incubation of Lactose with Streptococcus thermophilus and Lactobacillus Bulgaricus Bacteria

(29) In this study, the extent of lactose cleavage by lyophilized and resuspended cultures of Streptococcus thermophilus and Lactobacillus bulgaricus bacteria was investigated.

(30) The strain of Streptococcus thermophilus used was Streptococcus thermophilus ST-VC18 (deposition number: DSM 23319) and the strain of Lactobacillus bulgaricus used was Lactobacillus delbrueckii LB-VC18 (deposition number: DSM 23320).

(31) Lyophilisates of both cultures were resuspended in phosphate buffer, Ph 7.3, both separately and in a 1:1 mixture of both bacteria based on colony forming units (cfu). Additionally, investigations were made as to whether the lactase activity in these bacterial preparations is stable in an acidic milieu comparable to the stomach (simulated gastric fluid=SGF). In all cases lactose cleavage was tested in the presence of pancreatic enzymes (Kreon capsules containing, 10 000 units lipase, 8 000 units amylase and 600 units of pancreas protease according to Pharmacopoea Europea and sodium dodecylsulfate) from porcine pancreas containing active pancreas enzymes plus detergent to mimic bile activity (simulated intestinal fluid=SIF). The amount of glucose, as measured enzymatically by the glucose oxidase assay, was taken as a measure for lactose turnover. The glucose oxidase assay was used as described and as referred to in the Sigma Aldrich Chem Co. Data sheet glucose oxidase assay, 1996.

(32) Lyophilized bacteria (1 mg/ml) were resuspended in 1 ml phosphate buffer (0.1 mol/l). lactose (10 mmol/l) and Kreon in a concentration of 2 capsules per liter (recommended intake 1 to 2 capsules per meal) was added and the suspension was incubated for 10 and 30 minutes. In this experiment, the Lactobacillus bulgaricus incubation resulted in the liberation of 1.7 mmol/l glucose after 30 minutes and incubation with Streptococcus thermophilus yielded 2.8 mmol/l glucose, indicating turnovers of 17 and 28% respectively.

(33) In order to test the acid stability of the lactase activity, resuspended lyophilisates (1 mg/ml bacterial lyophilisate per ml suspension) were incubated with simulated gastric fluid, Ph 4.0, for 10, 30 and 60 minutes. The suspension was neutralized by addition of phosphate buffer to give a Ph of approximately 7.0, and then lactose (10 mmol/l) and Kreon were added and the lactase activity determined as described above (30 min incubation of bacterial suspension with lactose, i.e. identical conditions).

(34) Lactobacillus suspensions liberated 2.0 to 2.5 mmol/l glucose after 30 minutes and Streptococcus suspensions liberated 3 to 3.5 mmol/l glucose after this period. No difference was seen between the samples having been incubated for 10, 30 or 60 minutes with simulated gastric fluid.

(35) This experiment demonstrates that the lactase activity in both preparations is stable in a reconstituted stomach environment.

(36) In a third experiment Streptococcus thermophilus and Lactobacillus bulgaricus lyophilisates were mixed on a 1:1 basis (based on the numbers of colony forming units). For turnover measurements, 1 mg Streptococcus and 0.6 mg Lactobacillus bacteria were resuspended in 1 ml phosphate buffer and the resulting mixture was incubated with simulated intestinal fluid. After incubating this mixture for 30 minutes with 10 mmol/l lactose, 3.1 mmol glucose was detected.

(37) The same mixture was incubated for 30 minutes with simulated gastric fluid prior to lactase determination. After neutralisation and incubation for 30 minutes in simulated intestinal fluid with 10 mmol/l lactose, the concentration of glucose was found to be 3.15 mmol/l thereby demonstrating the stability of this mixture against acidic conditions.

(38) The experiments demonstrate the presence of lactase activity in lyophilized bacteria. This lactase activity is stable and active in simulated intestinal fluid, even after having been subjected to simulated gastric fluid before being tested in SIF. From these experiments it can be concluded that in vivo hydrolysis of lactose in the small intestine can be achieved by ingestion of the combination of both bacteria or by ingestion of each bacteria on its own and that there is no significant difference between the activities of the individual types of bacteria when used alone or when used in combination.

(39) The data provided in the examples above demonstrate that combinations of acid lactase and the bacteria Streptococcus thermophilus and/or Lactobacillus bulgaricus provide a more effective and more reliable improvement in lactose digestion in lactose-malabsorbing subjects than do compositions containing just the lactase or compositions containing just the bacteria. Furthermore, contrary to the teachings in the prior art documents discussed above, it is possible to present the bacteria in dried form and it is not necessary to coat the bacteria with an enteric coating in order to achieve the desired therapeutic effect.