Composition For Maintenance Of Bone Or Dental Health Or Treatment of Bone Or Dental Disorders
The present invention relates to a composition for maintenance of bone or dental health or treatment of bone or dental disorders, a method of preparing the composition; the composition for use as a medicament; use of the composition in the manufacture of a medicament or nutritional product; and a method of treatment of bone or dental disorders or maintenance of bone or dental health which comprises administering an effective amount of the composition.
Within the context of this specification the word "comprises" is taken to mean "includes, among other things". It is not intended to be construed as "consists of only".
Within the context of this specification RDA represents recommended daily amount, FCA represents fractional calcium absorption, OC represents osteocalcin, PINP represents pro-collagen type I N-terminal propeptide, APPase represents alkaline phosphatase, DPYR represents deoxypyridinoline, sXlaps and uXlaps represent serum and urinary crosslaps, respectively. FCA is determined after recovery of stable isotopes of calcium in urine samples. OC, APPase, PINP and sXlaps are detected in blood samples. DPYR and uXlaps are detected in urine samples. FCA depicts intestinal calcium absorption. OC, PINP and APPase are markers of bone formation. DPYR, s- and uXlaps are markers of bone resorption.
It is well known that minerals are key elements in several major physiological processes. Calcium is, for example, known to be of vital importance for the formation of bones and teeth, muscle contraction and the synthesis of hormones. Calcium is also an essential secondary messenger in several cellular activation pathways.
Minerals, of which the diet is the primary source, are assimilated by the body by crossing the intestinal mucosa to pass into the blood stream. The degree of assimilation (or of absorption) of minerals by the body depends both on their
solubility in the intestinal medium and on the capacity of the intestinal cells to assimilate and transfer them into the blood stream.
The location, efficiency and the mechanisms of calcium absorption along the intestine have been studied in rats and chickens for many years. However, for ethical and technical reason such studies have been limited in man.
One of the most widely studied aspects of mineral absorption is the bioavailability of the minerals in relation to the composition of the daily diet. However, many minerals which are highly bioavailable also cause organoleptic degradation in some products and may be unsuitable for use in such products. Furthermore, merely adding greater amounts of minerals often has a negative effect on the organoleptic quality of the product.
Rasic et al (Fermented Fresh Milk Product, vol 1, pi 14-115, 1978) reported that minerals contained in dairy products are assimilated better when the products are fermented. This effect is attributed to the presence of acids in fermented dairy products.
More recently, Yaeshima et al (Bulletin of International Dairy Fermentation, No. 313, 1996) showed an increase in the absorption of calcium in rats from a diet of calcium fortified whey when a combination of oligosaccharides and Bifidobacteria was consumed. However, Kot et al (J, Agric. Food Chem., 43_, 1276-1282, 1995) reported that Lactobacillus acidophilus naturally internalises Fe2+, and oxidises it to Fe3+ which is insoluble and more difficult to assimilate.
In addition to the problems described above, osteoporosis is recognised as a major public health problem in Western countries, especially in post menopausal women. Indeed, post-menopausal osteoporotic fractures affect about 1.5 million people per annum. About 300,000 new cases of osteoporotic hip, 650,000 vertebral and 200,000 distal forearm fractures are reported annually in the US. Mortality in the first year after hip fractures reaches 20% (Amer. J. Epidem. 1001-1005 (1993)). The estimated cost of treatment of these patients is in the region $6-$ 10 billion annually. About half the survivors are unable to walk unassisted and 25% are confined to long term care in a nursing home. This
problem is set to increase in view of the fact the life expectancy is increasing and a greater number of people are reaching old age.
Remarkably, it has now been found that a composition which comprises a probiotic bacterium and at least one component selected from the group consisting of calcium, calcium fortification compounds, calcium containing compounds and vitamin D has a positive affect on maintenance of bone or dental health or treatment of bone or dental disorders. This is unexpected because although the beneficial affect of calcium and/or vitamin D is well described it has now suprisingly been found that this affect may be increased in the presence of a probiotic bacterium.
Accordingly, in a first aspect the invention provides a composition which comprises a probiotic bacterium and at least one component selected from the group consisting of calcium and vitamin D.
In a second aspect, the invention provides a method of producing a composition having a probiotic bacterium and at least one component selected from the group consisting of calcium and vitamin D which comprises the steps of contacting a solution of calcium or a solution of vitamin D with a probiotic bacterium culture.
In a third aspect the invention provides a composition which comprises a probiotic bacterium and at least one component selected from the group consisting of calcium and vitamin D for use as a medicament.
In a fourth aspect the invention provides the use of a composition which comprises a probiotic bacterium and at least one component selected from the group consisting of calcium and vitamin D in the manufacture of a medicament or nutritive product for the treatment of a bone or dental disorder or maintenance of bone or dental health.
In a fifth aspect the invention provides a method of treatment of a bone or dental disorder or maintenance of bone or dental health which comprises administering an effective amount of a composition which comprises a probiotic bacterium and at least one component selected from the group consisting of calcium and vitamin D.
Preferably a composition according to an embodiment of the invention comprises a probiotic bacterium, calcium and vitamin D. More preferably a composition according to an embodiment of the invention also comprises phosphorus and/or magnesium ions and/or calcium citrate malate (CCM). CCM is well absorbed in man and provides the advantage of a high calcium intake or a higher bioavailability of calcium from the diet, even in diets deficient in calcium.
Preferably, an embodiment of the composition according to the invention comprises casein phosphopeptide (CPP). CPP provides the advantage of stimulating absorption of calcium intake from the diet, even where there is a deficiency of calcium in the diet.
Preferably, the bacterium is a lactic acid bacterium. More preferably, it is selected from the group which comprises Lactobacillus acidophilus,
Lactobacillus crispatus, Lactobacillus amylovorous, Lactobacillus gallinarum, Lactobacillus gas se and Lactobacillus johnsonii; Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus helveticus, Lactobacillus brevis; Lactobacillus fermentum; Lactobacillus plantarum; Lactobacillus casei particularly L. casei, Lactobacillus paracasei and L. rhamnosus; Lactobacillus delbrueckii particularly L. delbrueckii subsp lactis, L. delbrueckii subsp. delbrueckii and L. delbrueckii subsp. bulgaricus; and bifidobacteria particularly Bifidobacterium infantis, Bifidobacterium breve, Biβdobacterium adolescentis, Bifidobacterium lactis Biβdobacterium longum; and Leuconostoc mesenteroides particularly L. mesenteroides subsp cremoris. Most preferably the bacterium is Lactobacillus acidophilus LalO (ATCC 11975). An advantage provided by these bacteria is that they have a striking positive affect on enhancement of calcium absorption. Remarkably, the affects on bone formation and resorption are particularly striking when the composition comprises Lactobacillus acidophilus LalO (LalO).
Preferably a composition according to an embodiment of the invention comprises about 10 to about 10 cfu/ml or /g probiotic bacterium. More preferably it comprises about 10 cfu/ml probiotic bacterium.
Preferably a dose of about 250 to about 300g of a composition according to an embodiment of the invention is provided by two servings per day (eg. two yoghurts of about 125g to about 150g per day). More preferably the dose is about 250g per day ie two yoghurts of about 125g per day.
Preferably a composition according to an embodiment of the invention comprises about 15% to about 64% the RDA of calcium per 250g (RDA calcium is lOOOmg); ie about 75mg to about 320mg per 125g of the composition. More preferably it comprises about 50% of RDA calcium per 250g; ie 250mg per 125g of the composition.
Preferably, a composition according to an embodiment of the invention comprises about l μg to about 3.75μg vitamin D per 125g of the composition (yoghurt has been supplemented with 2.5 μg vitamin D/lOOg; thus, 2 such yoghurts of 125g provide more than 100% of RDA) . More preferably it comprises about 50% of RDA vitamin D per 250g (RDA vitamin D is 5μg); ie 1.25μg vitamin D per 125g of the composition.
Preferably, an embodiment of a composition according to the invention comprises nutrients/non-nutrients selected from the group which comprises:
Magnesium, calcium citrate malate or calcium citrate lactate or milk calcium, phosphorus, iron, iodine, zinc, copper, selenium;
Vitamin D, D3, A, Bl, B2, B6, B12, C, E, K, niacin, folic acid, pantothenic acid, biotin;
Prebiotics (eg fibres, cellulose);
Protein, casein phosphopeptide (CPP);
Isoflavones;
Bioactive peptide derived from milk eg TGF- , PTHrp.
Preferably the amount of magnesium, calcium citrate malate or calcium citrate lactate or milk calcium, phosphorus, iron, iodine, zinc, copper, selenium is
between about 15 and about 50% of RDA per 250g of the composition ie about 7.5 to about 25% RDA per 125g of the composition.
Preferably the amount of vitamin D, D3, A, Bl, B2, B6, B12, C, E, K, niacin, folic acid, pantothenic acid, biotin is between about 15 and about 33% of RDA per 125g of the composition.
Preferably the amount of prebiotics (eg fibres, cellulose)is about 6 to about 40g per 250g; ie about 3 to about 20g per 125g of the composition.
Preferably the amount of protein is about 2g to about 6g per 125g. More preferably it is about 4.4g per 125g of the composition.
Preferably the amount of casein phosphopeptide (CPP) is about 50 mg to about 2g per 250g of the composition; ie about 25mg to about lg per 125g of the composition.
Preferably the amount of isoflavones is about 25 to about 50 mg per 250g of the composition; ie about 12.5mg to about 25mg per 125g of the composition.
Preferably the amount of bioactive peptide derived from milk eg TGF- o r PTHrp is about 1.5 to about 2.5 ng per mg of protein.
Specific embodiments of the invention will now be described in further detail with reference to the accompanying drawings in which:
Figure 1 shows results of a study on the positive effect of LalO on intestinal calcium absorption (FCA) and two bone formation markers (OC and PINP) in adult men.
Figure 2 shows results of a study on the effect of LalO on a bone formation marker (PINP) in post menopausal women.
Figure 3 shows the results of a study on the beneficial effect of a milk calcium preparation on bone mineral density in prepubertal girls.
It is clear from the results shown in figure 1 that there is an improvement of calcium absorption and osteocalcin (bone formation) in the presence of lactic acid bacteria in men. In addition there is a trend for a decrease in deoxypyridinoline (DPYR) (bone resorption). In the light of the results obtained it may be concluded that a composition according to an embodiment of the invention may provide the benefit of maintenance of bone or dental health by positively affecting bone formation.
It is clear from the results shown in figure 2 that there is an improvement of PINP (bone formation) in women in the presence of lactic acid bacteria.
In the light of the results obtained it may be concluded that a composition according to an embodiment of the invention may provide the benefit of maintenance of bone or dental health by positively affecting bone formation.
Example 1
A composition according to an embodiment of the invention comprises about 250mg to about 320 mg calcium per 125g to 150g of the composition (which corresponds to about 30% calcium supplementation of a standard yoghurt; a standard yoghurt contains approx. 150 mg of calcium/ lOOg, i.e. 1.5 mg/g; a 30% supplementation would bring calcium concentration to 2 mg/g).
A composition in the form of a mixture of yoghurt, calcium derived from milk (Lactoval™ (DMV International)), vitamin D and LalO lactic acid bacteria was prepared by mixture of the components. 0.16% Lactoval™, 0.001% vitamin D3 (Roche, 100CWS/A), and an appropriate amount of LalO bacteria were added to 125g yoghurts in order to reach 200 mg calcium, 3 μg vitamin D and 109 CFU LalO per lOOg of yoghurt. Tests were conducted to check for positive affects on calcium absorption and bone formation.
Enrichment with Ca and Vitamin D was carried out during standardisation of milk. Both Ca and Vitamin D were mixed with the milk before pasteurisation and homogenisation.
Example 2
A composition was produced by simple mixture of yoghurt with LalO probiotic bacteria. The minimal cell count of LalO was 107/ml or g of yoghurt.
Example 3
Compositions which comprise the following were produced:
Milk calcium (Lactoval™ (DMV International)) in cakes;
Milk calcium (Lactoval™ (DMV International)) in biscuits;
Milk calcium (Lactoval™ (DMV International)) in chocolate bars;
Milk calcium (Lactoval™ (DMV International)) in fruit juices;
Milk calcium (Lactoval™ (DMV International)) in powdered drinking chocolate;
Milk calcium (Lactoval™ (DMV International)) in yoghurts;
In vitro data on LalO and Ca absorption and in vivo data on the effects of LalO on bone metabolism were obtained. It is presented in the figures.
The protocols for the in vivo studies were as follows:
In a first study, a two week wash-out period was followed by two weeks of test followed by three weeks of wash-out and two further weeks of test . 10 male and 21 female subjects were enrolled in a cross-over design so that each volunteer received yoghurt with LalO during one test period and yoghurt without LalO during the other test period. The study was designed to detect a 6% absolute increase in calcium absorption from the diet. One strain of LalO bacterium was used. The main parameters analysed were intestinal calcium absorption using a stable isotope technique; Lactic Acid Bacteria survival using recovery of LalO and influence on endogenous flora; bone formation using osteocalcin, alkaline phosphatase and PINP analysis (osteocalcin, alkaline phosphatase and PINP were
detected by immunoassays in blood samples); and bone resorption using serum and urinary crosslaps and DPYR analysis (serum and urinary crosslaps were detected in blood and urine samples, respectively; DPYR was detected by HPLC in urine samples.
The study on men revealed the following:
With regard to calcium absorption, there was a trend for an increase.
With regard to LalO recovery and influence on endogenous flora, there was a 50% recovery of LalO and a significant increase in total lactobacilli population.
With regard to bone turnover, there was a significant increase in osteocalcin, a bone formation marker, and a trend for a decrease in DPYR, a bone resorption marker.
The study on women revealed the following:
With regard to calcium absorption, variability of the results was slightly lower than expected, mean absorption was lower than in men.
With regard to LalO recovery, similar results to the study in men were achieved.
With regard to bone turnover, there was a significant increase in PINP, a bone formation marker
In a second study, 149 pre-pubertal girls received calcium-supplemented products, listed in Example 3, for 1 year. Bone mineral density (BMD) was determined by dual-energy X-ray absorptiometry at baseline, after 1 year of consumption, and after further 1 and 3.5 years without consumption of calcium- supplemented product.
The study on pre-pubertal girls revealed the following:
Girls receiving the calcium-supplemented products had a significantly higher gain in BMD.
Gains in BMD were maintained at least for 3.5 years.
In general the studies revealed the following results:
Milk Calcium
Helps increase bone mineral density during growth
LalO bacterium
Helps increase the amount of Ca absorbed Helps decrease bone loss Helps increase bone formation
LalO bacterium + milk Ca + vitamin D
Helps increase the amount of Ca absorbed Helps increase Ca absorption Helps decrease bone loss
Helps increase bone formation
Helps increase bone mineral density during growth
In the light of these results it was concluded that the results in men and in women show that bone formation may be significantly increased, and that the results in girls show that BMD may be significantly increased, if a diet including a composition according to the invention is administered.
It should be understood that various changes and modifications to the presently preferred embodiments described herein will be apparent to those skilled in the art. Such changes and modifications can be made without departing from the spirit and scope of the present invention and without diminishing its attendant advantages. It is therefore intended that such changes and modifications be covered by the appended claims.