FERMENTED DAIRY COMPOSITIONS AND METHODS OF PREPARING THE SAME

Abstract

The present invention relates to the use of L. acidophilus in the preparation of fermented dairy compositions having low levels of D-lactate.

Claims

1. A fermented dairy composition comprising L. acidophilus and up to 0.02% w/w D-lactate and having a pH equal to or lower than 5.

2. The fermented dairy composition according to claim 1, comprising L. acidophilus CNCM 1-2273.

3. The fermented dairy according to claim 1, further comprising S. thermophilus.

4. The fermented dairy composition according to claim 1, suitable to be administered to infants or young children.

5. The fermented dairy composition according to claim 1, comprising at least 810.sup.6 CFU/g L. acidophilus.

6. A process for the preparation of a fermented dairy composition comprising fermenting a mixture comprising a dairy composition and L. acidophilus to obtain a fermented dairy composition having up to 200 ppm d-lactate and a pH equal to or lower than 5.

7. The process according to claim 6, wherein said L. acidophilus is CNCM 1-2273.

8. A process for the preparation of a fermented dairy composition comprising fermenting a mixture comprising a dairy composition and L acidophilus CNCM 1-2273 and S. thermophilus to obtain a fermented dairy composition.

9. The process according to claim 6, wherein said fermented dairy composition comprises at least 810.sup.6 CFU/g L. acidophilus.

10. The process according to claim 6, wherein said mixture further comprises one or more of S. thermophilus and/or L. bulgaricus.

11. The process according to claim 6, wherein said fermentation is carried out at a temperature of less than about 42 C.

12. The process according to claim 6, further comprising storing said fermented dairy composition for at least 72 hours prior to consumption.

13. An inoculum comprising at least 10.sup.9 cfu per gram L. acidophilus CNCM 1-2273.

14. A mixture comprising an inoculum according to claim 13.

15. A mixture according to claim 14, further comprising at least one inoculum of Streptococcus thermophilus.

16. A mixture according to claim 14, further comprising at least one inoculum of Streptococcus thermophilus, one inoculum of Lactobacillus bulgaricus and optionally one or more additional inoculum of Lactococcus lactis or Lactobacillus acidophilus.

17. The fermented dairy composition according to claim 4, suitable to be administered starting from the age of 4 months.

Description

DESCRIPTION OF THE FIGURES

[0122] FIG. 1 shows milk acidification kinetics of L. acidophilus CNCM 1-2273 and control strain (strain X) together with S. thermophilus in milk. Time (in minutes) is provided on the x-axis, and pH is represented on the y-axis.

[0123] FIG. 2 shows milk acidification kinetics of L. acidophilus strains reference nos. 28, 30, 32, 43 and CNCM 1-2273 in milk (lait). Time (in minutes) is provided on the x-axis, and pH is represented on the y-axis.

[0124] FIG. 3 shows milk acidification kinetics of L. acidophilus strains reference nos. 28, 30, 32, 43 and CNCM 1-2273 in milk (lait) supplemented with 20 g/l glucose. Time (in minutes) is provided on the x-axis, and pH is represented on the y-axis.

[0125] FIG. 4 shows milk acidification kinetics of L. acidophilus strains reference nos. 28, 30, 32, 43 and CNCM 1-2273 in milk (lait) supplemented with 0.5 g/l N3 peptide (Vitalarmor 950). Time (in minutes) is provided on the x-axis, and pH is represented on the y-axis.

[0126] FIG. 5 shows milk acidification kinetics of L. acidophilus strains reference nos. 28, 30, 32, 43 and CNCM 1-2273 in milk (lait) supplemented with 2 g/l yeast extract (YE). Time (in minutes) is provided on the x-axis, and pH is represented on the y-axis.

EXAMPLES

Example 1: Screening of Strains

[0127] A screening of 5 strains of L. acidophilus was carried out to identify strains that were suitable for the preparation of fermented milk products based on their milk acidification properties. The test strains for screening purposes were provided by Danone: L. acidophilus strains reference nos. 28, 30, 32, 43 and CNCM 1-2273. The test strains were tested in milk (FIG. 2), milk supplemented with 20 g/l glucose (FIG. 3), milk supplemented with 0.5 g/l N3 peptide (Vitalarmor 950) (FIG. 4), and milk supplemented with 2 g/l yeast extract (FIG. 5).

[0128] Fermented milk test products were prepared by combining whole cow milk (3.2% protein content; autoclaved at 120 C. 15 minutes for sterilization) with inoculum (pure strains of L. acidophilus) and fermenting at 37 C. The inoculum of L. acidophilus was in thawed frozen form provided from the Danone bacterial strain collection.

[0129] As can be seen from FIGS. 2-5, strain CNCM 1-2273 had above-average milk acidification kinetics. This was unexpected in view of the present invention in WO 2010/058294 discussed above.

Example 2: Fermented Milk Product Preparation

[0130] Fermented milk test products were prepared by combining cow milk having 3% protein and 3.4% fat with inoculum and fermenting.

[0131] Bacterial strains were provided in frozen form. The inoculum comprised a S. thermophilus inoculum together with one of the test L. acidophilus strains inoculum.

[0132] The S. thermophilus used F-DVS ST-BODY-4 is a commercially available strain from Chr. Hansen.

[0133] A commercially available L. acidophilus strain (Control strain) was used as a control. This strain is considered as having acceptable levels of D-lactate production as demonstrated by long-standing commercial use with no associated consumer illness or injury as well as FDA GRAS and EFSA QPS status. The inoculum of Control strain contained 610.sup.10 CFU/g.

[0134] Strain CNCM 1-2273 was tested in liquid frozen (thawed) format: [0135] inoculum ino 1 was prepared by propagation on a medium containing milk-derived components
inoculum ino 2 was prepared by propagation on a medium of milk-derived components. The inoculums had 3.110.sup.10 CFU/g (ino 1) and 2.610.sup.10 CFU/g (ino 2) in frozen format.

[0136] The strains were inoculated in the milk in the following volumes to ensure comparable amounts of the strains of L. acidophilus:

S. thermophilus 0.03% v/v
L. acidophilus Control strain 0.02% v/v
L. acidophilus CNCM 1-2273 0.04% v/v

[0137] CNCM 1-2273 was also tested with the addition of 0.008% v/v yeast extract in the milk mixture.

[0138] Fermentation was carried out at 42 C. and monitored using a CINAC pH probe. The resultant fermented milk was cooled and stored at 10 C. for determination of strain viability and D-lactate content after 3 days of storage.

[0139] Results

[0140] In all samples, D-lactate levels were detectable and lower than the target of 200 ppm. FIG. 1 provides the acidification kinetics of the test products.

TABLE-US-00001 TABLE 1 Characteristics of fermented milk products L. acidophilus fermentation Inoculation S. thermophilus L. acidophilus time population count Day 3 count Day 3 pH [H] [CFU/g] [CFU/g] [CFU/g] Control strain 5.04 08:20 1.20E+07 3.51E+07 2.68E+06 CNCM I-2273 4.69 07:10 1.24E+07 1.45E+08 1.84E+07 Frozen ino 1 CNCM I-2273 4.69 05:58 1.24E+07 1.22E+08 1.78E+07 Frozen ino 1 + yeast extract CNCM I-2273 4.68 06:43 1.04E+07 1.28E+08 1.31E+07 Frozen ino 2+ CNCM I-2273 4.68 06:08 1.04E+07 1.40E+09 1.65E+08 Frozen ino 2 + yeast extract

[0141] As can be seen from the above results: CNCM 1-2273 produces acceptable levels of D-lactate, as does Control strain but at approximately one log higher population (CFU). CNCM 1-2273 growth in milk (CFU count) was higher than the commercially used strain, especially when supplemented with yeast extract. This indicates that the D-lactate per CFU production is lower in CNCM 1-2273 than Control strain.

[0142] CNCM 1-2273 population was comparable between ino 1 and 2, however, some difference in D-lactate levels observed between different propagation conditions.

[0143] A trend towards symbiosis between CNCM 1-2273 and S. thermophilus can be observed as ST-BODY-4's growth is impacted by a higher count of CNCM 1-2273 (+1 log for ST-BODY-4 in the last sample) and a faster fermentation time than was required for Control strain.

[0144] D-lactate levels in all the fermented milk products was comparable. However, taking into account that the population of CNCM 1-2273 was significantly higher, it can be concluded that higher probiotic content can be achieved in fermented milk products with CNCM 1-2273, while ensuring acceptable D-lactate levels or that the probiotic count can be reducedwith an expected reduction in D-lactate levels.

Example 3: Confirmation of CFU/D-Lactate Production

[0145] Fermented milks prepared according to Example 2. Multiple fermentations were carried out using the test (CNCM 1-2273) and control strain both in combination with the ST-BODY-4 and in an alternative culture with an alternative ST-Body commercially available strain. Fermentation was carried out until bacterial counts in the ranges of 10.sup.6-10.sup.8 were achieved (at a pH of about 4.7) and D-lactate productions was measured.

[0146] A CFU/D-lactate relationship was confirmed, the higher the CFU the higher the D-lactate. It was also confirmed that D-lactate production of CNCM 1-2273 was significantly lower per CFU compared to control strain.

[0147] Fermented milk could be produced with a CFU of up to 110.sup.8 CFU/g, more preferably 2.510.sup.7 CFU/g when using CNCM 1-2273, however if using control strain the maximum CFU count achievable was 810.sup.6 CFU/g.

[0148] Thus a particularly interesting target range for having a high cfu/g probiotic product with good D-lactate levels would be within the range of at least 810.sup.6 cfu/g and up to 110.sup.8 cfu/g. However, to ensure that margin of error is allowed the upper and limit may be set more conservatively (e.g. not more than 2.5, 5 or 7.510.sup.7 CFU/g L. acidophilus as the upper limit; at least 8.1, 8.3, 8.5 or 8.710.sup.6 or 110.sup.7 CFU/g as the lower limit).