CN115531424A - Probiotic drops - Google Patents
Probiotic drops Download PDFInfo
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- CN115531424A CN115531424A CN202110759489.0A CN202110759489A CN115531424A CN 115531424 A CN115531424 A CN 115531424A CN 202110759489 A CN202110759489 A CN 202110759489A CN 115531424 A CN115531424 A CN 115531424A
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- probiotic
- drops
- lactobacillus
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Abstract
The present invention relates to probiotic drops. The invention discloses a probiotic drop, wherein the drop comprises the following components by weight based on the total weight of the drop: 1-15wt% of probiotic bacteria; 31-50wt% sunflower seed oil; 31-50wt% olive oil; and 17-30wt% of triglyceride.
Description
Technical Field
The invention relates to a probiotic drop (probiotic drop), wherein the drop comprises, based on the total weight of the drop: 1-15wt% of probiotic bacteria; 31-50wt% sunflower oil (sunflower oil); 31-50wt% olive oil (olive oil); and 17-30wt% of triglyceride (triglyceride).
Background
Probiotics selectively stimulate the growth of native bacteria (native bacteria) in the gut, alter the composition, quantity or nature of the gut microbiota (intestinal microflora), and when administered in appropriate amounts, confer a health benefit (health benefit) to the host. Microorganisms currently known for use as probiotics include: species such as Lactobacillus, lactococcus, pediococcus, enterococcus, streptococcus, bifidobacterium, bacillus, and Leuconostoc.
Many studies have shown that probiotics have the effects of inhibiting the growth of gastrointestinal pathogens, alleviating lactose intolerance (lactose intemperance), preventing and/or treating gastrointestinal diseases (gastrointestinal disease), immunomodulation (immunomodulation), anti-bacterial (anti-bacterial), anti-cancer (anti-cancer) and blood pressure lowering (anti-hypertension).
However, during the preparation of probiotic preparations (probiotic preparations), the survival time of the probiotics in the probiotic preparations is affected by the type of the selected preparation formulation and the storage environment (e.g., temperature and humidity), and thus the number of bacteria required for the probiotics preparation to exert effect cannot be maintained. Therefore, how to maintain the number of probiotics in the probiotic preparation has become an objective of the relevant researchers in this field.
In recent years, researchers of interest have attempted to increase the survival rate of probiotics by dispersing the probiotics in lipid solvents. For example, CN 105707897A discloses a probiotic lipid suspension drop, which comprises probiotic powder, vegetable oil, medium chain glyceride (medium chain glyceride), sucrose monoglyceride (sucrose monoglyceride) and vitamin E (vitamin E). In the example of this piece of chinese patent publication, sunflower oil (sunflower oil) was used to formulate lipid suspension drops of Lactobacillus reuteri and Lactobacillus plantarum, respectively, and demonstrate that the bacterial count of the probiotic bacteria could be maintained.
Despite the above-mentioned reports, there is still a need in the art to develop a probiotic preparation that can effectively improve the stability of probiotics for industrial needs.
Disclosure of Invention
In the present invention, the applicant found that a probiotic drop (probiotic drop) containing probiotics, sunflower oil (sunflower oil), olive oil (olive oil) and triglyceride (triglyceride) can effectively improve the stability of the probiotics, and further can maintain the activity of the probiotics under long-term storage.
Accordingly, the present invention provides a probiotic drop wherein the drop comprises, based on the total weight of the drop:
1-15wt% of probiotic bacteria;
31-50wt% sunflower seed oil;
31-50wt% olive oil; and
17-30wt% of triglyceride.
Preferably, the sunflower oil is present in substantially the same amount as olive oil.
Preferably, the drop further comprises: 0.1-1.5wt% sucrose monoglyceride; and 0.01-0.05wt% of vitamin E.
Preferably, the probiotic is selected from the group consisting of: lactobacillus species (Lactobacillus spp.), lactococcus species (Lactococcus spp.), pediococcus species (Pediococcus spp.), enterococcus species (Enterococcus spp.), streptococcus species (Streptococcus spp.), bifidobacterium species (Bifidobacterium spp.), bacillus spp., leuconostoc spp., and combinations thereof.
More preferably, the probiotic is selected from the group consisting of: lactobacillus rhamnosus (Lactobacillus), bifidobacterium animalis subsp.
Preferably, the drop further comprises a desiccant.
Drawings
The above and other objects, features and advantages of the present invention will become apparent upon consideration of the following detailed description and preferred embodiments thereof, taken in conjunction with the accompanying drawings, in which:
FIG. 1 shows the bacterial concentration of various drops containing Lactobacillus rhamnosus MP108 as a function of time when stored at 37 ℃ wherein drops 1 to 7 have a formulation as shown in Table 2 below; and
figure 2 shows the bacterial concentration of various drops containing bifidobacterium animalis subsp lactis CP-9 as a function of time when stored at 37 c, where drops 1 to 7 generally have a formulation as shown in table 2 below, with the difference that: the bacterial powder of bifidobacterium animalis subsp lactis CP-9 is used for replacing the bacterial powder of lactobacillus rhamnosus MP108.
Detailed Description
For the purposes of this specification, it will be clearly understood that: the word "comprising" means "including but not limited to", and the word "comprising" has a corresponding meaning.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. One skilled in the art will recognize many methods and materials similar or equivalent to those described herein, which can be used in the practice of the present invention. Of course, the present invention is in no way limited to the methods and materials described.
The present invention provides a probiotic drop (probiotic drop), wherein the drop comprises, based on the total weight of the drop:
1-15wt% of probiotic bacteria;
31-50wt% sunflower oil (sunflower oil);
31-50wt% olive oil (olive oil); and
17-30wt% of triglyceride (triglyceride).
According to the invention, the sunflower oil has substantially the same content as olive oil.
As used herein, the term "substantially identical" means having equal amounts (equivalents) or differing by less than 1.5 wt%. Preferably, the sunflower seed oil has exactly the same content as the olive oil.
According to the invention, the drops have a concentration of bacteria in the range of 8.5 to 10.6log CFU/mL of probiotic bacteria. In a preferred embodiment of the invention, the drops have a concentration of bacteria of 10.4log CFU/mL of probiotic bacteria. In another preferred embodiment of the invention, the drops have a concentration of bacteria of 10.6log CFU/mL.
According to the invention, the probiotic bacteria may be natural materials or processed products (e.g. freeze-dried or spray-dried products). Preferably, the probiotic is a processed product. More preferably, the probiotic is a powder of bacteria obtained by freeze-drying followed by grinding.
According to the invention, the probiotic may be selected from the group consisting of: lactobacillus species (Lactobacillus spp.), lactococcus species (Lactococcus spp.), pediococcus species (Pediococcus spp.), enterococcus species (Enterococcus spp.), streptococcus species (Streptococcus spp.), bifidobacterium species (Bifidobacterium spp.), bacillus spp., leuconostoc spp., and combinations thereof.
Preferably, the probiotic is a lactobacillus species selected from the group consisting of: lactobacillus rhamnosus (Lactobacillus rhamnous), lactobacillus plantarum (Lactobacillus plantarum), lactobacillus acidophilus (Lactobacillus acidophilus), lactobacillus casei (Lactobacillus casei), lactobacillus paracasei (Lactobacillus paracasei), lactobacillus reuteri (Lactobacillus reuteri), lactobacillus bulgaricus (Lactobacillus bulgaricus), dai Baishi Lactobacillus delbrueckii), lactobacillus johnsonii (Lactobacillus johnsonii), lactobacillus helveticus (Lactobacillus helveticus), lactobacillus gasseri (Lactobacillus helveticus), lactobacillus fermentum (Lactobacillus fermentum), and combinations thereof. In a preferred embodiment of the invention, the probiotic is lactobacillus rhamnosus MP108.
Preferably, the probiotic is a bifidobacterium species selected from the group consisting of: bifidobacterium animalis subsp. lactis, bifidobacterium bifidum (Bifidobacterium bifidum), bifidobacterium lactis (Bifidobacterium lactis), bifidobacterium longum (Bifidobacterium longum) [ e.g., bifidobacterium longum (Bifidobacterium longum subsp. Longum) ], bifidobacterium breve (Bifidobacterium breve), bifidobacterium adolescentis (Bifidobacterium adolescentis), bifidobacterium infantis (Bifidobacterium infantis), and combinations thereof. In a preferred embodiment of the invention, the probiotic is Bifidobacterium animalis subsp lactis CP-9.
Alternatively, the probiotic may also be selected from the group consisting of: lactococcus lactis (Lactococcus lactis), streptococcus thermophilus (Streptococcus thermophilus), bacillus coagulans (Bacillus coagulousns), and combinations thereof.
According to the invention, the drop further comprises, based on the total weight of the drop:
0.1-1.5wt% of alcohol ester emulsifier (alcohol ester type emulsifying agent); and
0.01-0.05wt% of vitamin E (vitamin E).
Preferably, the alcohol ester emulsifier is selected from the group consisting of: sucrose glyceride (sucrose), polysorbate 80 (polysorbate 80), polyglycerol fatty acid esters (polyglycerol fatty acid esters), and combinations thereof. In a preferred embodiment of the present invention, the alcohol ester emulsifier is sucrose monoglyceride (sucrose).
According to the present invention, the drop may further comprise a drying agent (desiccant).
The desiccants suitable for use in the present invention may be commercially available products including, but not limited to: a dry TABLET (obtained from Aptar CSP Technologies under the product model number M-3061-133ACTIVE TABLET) and a dry film (obtained from CSP Technologies under the product model number M-0026 ACTIVE-Strip) TM )。
According to the invention, the drops may further comprise other pharmaceutically acceptable additives (additives) which are widely used in pharmaceutical manufacturing technology, such as preservatives (preservative), antioxidants (antioxidant), absorption enhancers (absorption enhancer) and stabilisers (stabilizing agent). The selection and amounts of such additives are within the skill and routine skill of those skilled in the art.
In a preferred embodiment of the invention, the drops comprise, based on the total weight of the drops: 7wt% of probiotics, 31.94wt% of sunflower oil, 31.94wt% of olive oil, 28.9wt% of triglyceride, 0.2wt% of sucrose monoglyceride and 0.02wt% of vitamin E.
In another preferred embodiment of the invention, the drops comprise, based on the total weight of the drops: 7wt% probiotics, 47.94wt% sunflower oil, 15.94wt% olive oil, 28.9wt% triglycerides, 0.2wt% sucrose monoglyceride and 0.02wt% vitamin E.
In yet another preferred embodiment of the invention, the drops comprise, based on the total weight of the drops: 7wt% probiotic bacteria, 15.94wt% sunflower oil, 47.94wt% olive oil, 28.9wt% triglycerides, 0.2wt% sucrose monoglyceride and 0.02wt% vitamin E.
The probiotic drops according to the invention may be administered orally (orally) in a single dose or in divided doses.
The invention will be further described with respect to the following examples, but it should be understood that these examples are for illustrative purposes only and should not be construed as limiting the practice of the invention.
< example >
General experimental materials:
1. probiotic bacterial strains:
the probiotic strains used in the following examples for efficacy evaluation have been deposited at the Center for biological resource conservation and Research Center (BCRC) of Food Industry Research and Development Institute (FIRDI) in taiwan, food Industry, research and Development Institute (300 new bamboo Food road 331, taiwan), and are publicly available. In addition, the probiotic strains are also deposited in the German Collection of microorganisms (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, DSMZ) and the Chinese Collection of Type cultures (CCTCC), respectively, according to the Budapest Treaty. For clarity, the information (including the academic name, the related origin, the accession number and the accession date) of each probiotic strain is integrated in the following Table 1.
TABLE 1 information on individual probiotic strains
2. Preparation of bacterial powders of various probiotic strains:
first, 2 probiotic strains described in item 1 above were inoculated into MRS broth (MRS broth) supplemented with 0.05% cysteine (cysteine), respectively, and cultured at 37 ℃ for 24 hours to activate the strains. Then, the activated strains were inoculated in MRS broth at an inoculum size of 2% (v/v), respectively, and cultured under an anaerobic condition at 37 ℃ over night. The resulting cultures were centrifuged at 3,000rpm for 10 minutes at 4 ℃ after counting the number of bacteria in a plate counting medium, followed by removal of the supernatant, and the resulting pellets (pellets) were each taken to mix with a freeze-drying protecting agent (freeze-drying protecting agent). Then, the resulting mixture was put in a liquid nitrogen bucket at-196 ℃ for quick freezing, and then dried. Then, the mixture was ground by a pulverizer (model number Comil U20, manufactured by Quadro) and then sieved by a 40-mesh sieve, thereby obtaining powders of Lactobacillus rhamnosus MP108 and Bifidobacterium animalis subsp.lactis CP-9 (each having 5.7X 10. Sup. Times. 10. Sup. Respectively) 11 CFU/g and 3.7X 10 11 Bacterial concentration of CFU/g).
3. The 3 vegetable oils sunflower oil (sunflower oil), olive oil (olive oil) and peanut oil (peanout oil) used in the examples below were obtained from Yu Guige food, soy and Taishan industries, respectively.
4. The triglycerides (triglycerides), sucrose monoglycerides (sucrose esters) and vitamin E (vitamin E) used in the examples below were obtained from Yuba corporation, zhengxing food additives, and Xinjiang corporation, respectively.
Example 1 preparation of drops (drop) containing probiotic bacteria
In this example, the applicant prepared 7 drops containing lactobacillus rhamnosus MP108 (i.e. drop 1 to drop 7 of MP 108) according to the formulation shown in table 2 below. The preparation method of various drops is carried out according to the following steps: first, vegetable oil, triglyceride, sucrose monoglyceride and vitamin E were mixed and stirred until emulsification (emulisification), then bacterial powder was slowly added and stirred well to obtain drops containing probiotic bacteria (bacteria concentration of 10.6log CFU/mL), each drop having a volume of 8mL and a total weight of 7.6g.
TABLE 2 formulation of drops containing probiotics
In addition, 7 drops containing Bifidobacterium animalis Lactobacillus subsp CP-9 (i.e., drops 1 to 7 of CP-9) were prepared substantially according to the formulation shown in Table 2 above, except that: the bacterial powder of bifidobacterium animalis subsp lactis CP-9 is used for replacing the bacterial powder of lactobacillus rhamnosus MP108, and the drops contain probiotics with the bacterial concentration of 10.4log CFU/mL.
Example 2 Effect of probiotic-containing drops made with different vegetable oils on probiotic stability
In this example, in order to investigate the effectiveness of probiotic containing drops prepared using different vegetable oils in improving the stability of probiotics, the applicant carried out stability tests (stability test) on 14 probiotic containing drops obtained according to example 1 above.
The experimental method comprises the following steps:
the 14 probiotic bacteria-containing drops were stored in a constant temperature and humidity chamber (ATH-120, available from grand precision industries, ltd.) for 28 days at a temperature set at 37 ℃, and 2mL of each of the 14 probiotic bacteria-containing drops were diluted before starting storage (i.e., day 0) and at 14 and 28 days after starting storage by adding 18mL of 0.1% peptone diluent (available from Merck), followed by counting the number of bacteria in a plate count medium.
As a result:
figure 1 shows the bacterial concentration of various drops containing lactobacillus rhamnosus MP108 as a function of time when stored at 37 ℃. As can be seen from fig. 1, the number of bacteria contained in drops 1 to 7 gradually decreases with increasing storage time, wherein the number of bacteria contained in drops 3, 4 and 5 decreases by a significantly smaller amount than those contained in drops 1, 2, 6 and 7. In particular, the bacterial concentration of drops 3, 4 and 5 was also maintained above 8.5log CFU/mL after 28 days of storage, with the highest bacterial concentration of drops 3.
Figure 2 shows the bacterial concentration of various drops containing Bifidobacterium animalis Lactobacillus subsp CP-9 as a function of time when stored at 37 ℃. As can be seen from fig. 2, the number of bacteria contained in drops 1, 2, 6 and 7 decreased with increasing storage time, whereas the number of bacteria contained in drops 3, 4 and 5 decreased slowly and the concentration of bacteria was maintained above 9log CFU/mL, with the highest concentration of bacteria in drops 3.
The results of these experiments show that: compared with the single use of the sunflower seed oil and the olive oil or the combination of the sunflower seed oil and the olive oil and the peanut oil, the drop prepared by the combined use of the sunflower seed oil and the olive oil can show more excellent effect on improving the stability of probiotics.
Example 3 Effect of drops containing probiotic bacteria with addition of drying agent (drying agent) on the stability of probiotic bacteria
In this example, to see if the probiotic bacteria-containing drops of the present invention still have the above excellent effect and the desiccant further enhances the effect in the case of oxidation of fats and oils which may occur when the consumer contacts the air during daily use, the applicant carried out the following experiment on the MP108 drop 3 obtained in example 1 above and additionally prepared a drop with a desiccant for comparison.
The experimental method comprises the following steps:
first, a drop with a desiccant added (hereinafter referred to as drop 3-1) was prepared substantially according to the formulation of drop 3 shown in Table 2 above, and a dry tablet (obtained from Aptar CSP Technologies under the product model number M-3061-133ACTIVE TABLET,8.2mm (width) 17.7mm (length) 2.5mm (height) was additionally added.
Then, drop 3 and drop 3-1 were stored in a constant temperature and humidity machine set at 25 ℃ for 168 days, and during the storage period, the drops were exposed to air for 30 seconds each day to simulate the situation of use of the consumer, and 2mL of each of the 2 drops were diluted by adding 18mL of 0.1% peptone diluent before starting the storage (i.e., day 0) and at the 112 th, 140 th and 168 th days after starting the storage, followed by counting the number of bacteria in a plate count medium.
As a result:
table 3 below shows the change in bacterial concentration over time between drop 3 and drop 3-1 when stored at 25 ℃.
TABLE 3 bacterial concentrations of drops 3 and 3-1
As can be seen from table 3, both drop 3 and drop 3-1 contained bacteria at a count above 9.4log CFU/mL during storage over a period of 112 days to 168 days, and in particular, drop 3-1 maintained a bacterial concentration of 10log CFU/mL after storage over 168 days. The results of this experiment show that: the probiotic bacteria-containing drops of the present invention are still effective in maintaining the stability of the probiotic bacteria in daily use, and the addition of a drying agent to the probiotic bacteria-containing drops of the present invention can achieve better effectiveness.
Combining the above experimental results, the applicant believes that: the drop prepared by combining the sunflower seed oil and the olive oil can effectively improve the stability of probiotics, has the effect of maintaining the activity of the probiotics under long-term storage, and can further add a drying agent to achieve more remarkable excellent effect.
All patents and publications cited in this specification are herein incorporated by reference in their entirety. In case of conflict, the present detailed description, including definitions, will control.
While the invention has been described with reference to the specific embodiments described above, it will be apparent that many modifications and variations can be made without departing from the scope and spirit of the invention. It is the intention, therefore, to be limited only as indicated by the claims appended hereto.
Claims (6)
1. A probiotic drop, characterized by: the drops comprise, based on the total weight of the drops:
1-15wt% of probiotic bacteria;
31-50wt% sunflower seed oil;
31-50wt% olive oil; and
17-30wt% of triglyceride.
2. Probiotic drops according to claim 1, characterized in that: the sunflower oil has substantially the same content as olive oil.
3. Probiotic drops according to claim 1, characterized in that: said drops further comprising:
0.1-1.5wt% sucrose monoglyceride; and
0.01-0.05wt% of vitamin E.
4. Probiotic drops according to claim 1, characterized in that: the probiotic is selected from the group consisting of: lactobacillus spp, lactococcus spp, pediococcus spp, enterococcus spp, streptococcus spp, bifidobacterium spp, bacillus spp, leuconostoc spp and combinations thereof.
5. Probiotic drops according to claim 4, characterized in that: the probiotic is selected from the group consisting of: lactobacillus rhamnosus (Lactobacillus), bifidobacterium animalis subsp.
6. Probiotic drops according to claim 1, characterized in that: the drops further comprise a desiccant.
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