CN101914518A - Method for preparing Lactobacillus casei microcapsule bacterial powder by spray drying - Google Patents
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Abstract
The invention discloses a method for preparing Lactobacillus casei microcapsule bacterial powder by spray drying, and aims to provide a method which can enhance drying resistance and product viable count of Lactobacillus casei by using various activity protection techniques and can realize large-scale production. The method comprises the following steps: Lactobacillus casei viable bacteria culture obtained by high density culture adopts pre-culturing or heat activation treatment as the metabolic regulation means, and the protector is made from trehalose, sucrose, sodium borate, sodium glutamate or glutathione and vitamin C; and the mixture of skimmed milk powder and beta-cyclodextrin is used as a wall material, a Lactobacillus casei thallus suspension containing the protector is used as a core material, and spray drying is carried out to obtain the Lactobacillus casei microcapsule bacterial powder. In the spray drying process, the charging temperature is 25 DEG C, the inlet air temperature is 130 DEG C, and the pressure of an atomizer is 150 kPa. The survival rate of the bacterial powder is higher than 60%, and the viable count is more than 1010 per g.
Description
Technical field
The present invention relates to a kind of method of utilizing spray drying method for preparation Lactobacillus casei microcapsule bacterium powder.
Background technology
Spray drying technology has that equipment is simple, cost is low, be easy to promote, help advantage such as large scale continuous prod, and particularly medicine, foodstuffs industry are widely used at commercial field.Spray drying technology is beneficial to probiotic bacterium scale operation, transportation and storage.But in the spray-drying process, microorganism will experience high temperature and exsiccant pressure, and cell injury mainly is because high temperature and drying and dehydrating.Temperature has the greatest impact to microbial survival, high temperature particularly, and high temperature can maintain the enzyme system of vital movement by destroy microorganisms, makes its active reduction or forfeiture.Because the high molecular hydrogen bonds of important biomolecule such as protein, nucleic acid and enzyme system are damaged, and cause bacterium protein to solidify sex change, degraded sex change inactivation takes place in nucleic acid, thereby destroys the composition of cell; The lipoids composition forms minimum hole on the heat of solution cytolemma, and entocyte is leaked, and also cell membrane, cell walls, tenuigenin and nucleic acid cause damage, thereby cause death.Spray-drying process causes that cell dries out gradually; and the cell dehydration may cause the irreversible change of cytolemma, protein structure and functional completeness; these all make the survival rate and the viable count of the probiotics bacterial powder that obtains by spray drying process receive certain influence; therefore, protect these basic functions and structure for cell survival with keep it functional most important.
Summary of the invention
The present invention is in order to overcome weak point of the prior art; provide a kind of combination to improve the resist drying ability of lactobacterium casei by the various active resist technology; improve the product viable count; and; the method for preparing Lactobacillus casei microcapsule bacterium powder simple to operate, continuous, quick, as can to accomplish scale production.
The present invention is achieved through the following technical solutions:
A kind of method of utilizing spray drying method for preparation Lactobacillus casei microcapsule bacterium powder is characterized in that, comprises the steps:
(1) high-concentration culturing: the lactobacterium casei yeast culture thing after will activating is that 1% inoculum size is seeded to the MRS liquid nutrient medium with volume fraction, cultivates 24 hours down at 37 ℃, makes the maximum viable count of cultivating reach 10
10Individual/more than the ml;
(2) adopt pre-cultivation or heat shock to handle as the metabolism regulating measure:
Described pre-cultural method is: the lactobacterium casei viable bacteria culture that step (1) high-density culture is obtained cooled off 10-20 minute in 4 ℃ of water-baths, centrifugal collection lactobacterium casei bacterial sediment thing, afterwards the protective material annex solution is joined in the lactobacterium casei of the collection bacterial sediment thing after centrifugal by aseptic technique, shake to make in 5 minutes and suspend evenly, 4 ℃ leave standstill 1h, afterwards, continue down to cultivate 2 hours, obtain containing protectant lactobacterium casei thallus suspension liquid in 37 ℃;
Described heat shock treatment process is: the lactobacterium casei viable bacteria culture that step (1) high-density culture is obtained heats 45min in heat-shock temperature is 50 ℃ water-bath, afterwards, cultivate 1h, centrifugal collection lactobacterium casei bacterial sediment things down at 37 ℃; The protective material annex solution is joined in the lactobacterium casei of the collection bacterial sediment thing after centrifugal by aseptic technique, shake to make in 5 minutes and suspend evenly, 4 ℃ leave standstill 1h, obtain containing protectant lactobacterium casei thallus suspension liquid;
Wherein, described protective material is by trehalose, sucrose, Sodium Tetraborate, Sodium Glutamate or gsh, vitamins C is formed, the add-on of the per-cent meter of the bacterial sediment thing weight after the lactobacterium casei that obtains according to collection is centrifugal is: the 0.6-1.2% trehalose, 2-8% sucrose, the 0.01-0.03% Sodium Tetraborate, 0.8-1.5% Sodium Glutamate or 0.8-1.6% gsh, the 0.02-1.0% vitamins C, and obtain: earlier with Sodium Tetraborate by following method, Sodium Glutamate or gsh mix, the water dissolution of the bacterial sediment thing identical weight after the lactobacterium casei that adds and collect is centrifugal, add trehalose at last, sucrose and vitamins C, mix, sterilized 15 minutes for 121 ℃ afterwards, make the protective material annex solution;
When the big synergism of needs, also need to add N.F,USP MANNITOL and soybean lecithin; the add-on of the per-cent meter of the bacterial sediment thing weight after the lactobacterium casei that obtains according to collection is centrifugal is: 1-4% N.F,USP MANNITOL; the 4-8% soybean lecithin; protective material obtains by following method: earlier with Sodium Tetraborate; Sodium Glutamate or gsh; N.F,USP MANNITOL and soybean lecithin mix; the water dissolution of the bacterial sediment thing identical weight after the lactobacterium casei that adds and collect is centrifugal; add trehalose at last; sucrose and vitamins C; mix; sterilized 15 minutes for 121 ℃ afterwards, make the protective material annex solution.
The add-on of the per-cent meter of the bacterial sediment thing weight after the lactobacterium casei that each component in the protective material obtains according to collection is centrifugal is: the 0.6-1.2% trehalose; 2-8% sucrose; the 0.01-0.03% Sodium Tetraborate; the 0.8-1.5% Sodium Glutamate; 1-4% N.F,USP MANNITOL; the 0.8-1.6% gsh; 4-8% soybean lecithin and 0.02-1.0% vitamins C; and obtain: earlier with Sodium Tetraborate by following method; Sodium Glutamate; N.F,USP MANNITOL; gsh and soybean lecithin mix; the water dissolution of the bacterial sediment thing identical weight after the lactobacterium casei that adds and collect is centrifugal; add trehalose at last; sucrose and vitamins C; mix; sterilized 15 minutes for 121 ℃ afterwards, make the protective material annex solution.
(3) preparation Lactobacillus casei microcapsule:
Select for use in the mixture of mixture, skim-milk and gum arabic of mixture, skim-milk and gelatin of skim-milk and beta-cyclodextrin any as the wall material, contain protectant lactobacterium casei thallus suspension liquid as core;
Add the water dissolution of 3 times of wall material weight in the wall material, the 8000r/min homogeneous is 5 minutes afterwards, sterilizes 15 minutes, and is cooled to 40 ℃ for 121 ℃; Add the Tween 80 that contains protectant lactobacterium casei thallus suspension liquid and wall material solution weight 0.1% again, 5000r/min second homogenate 10 minutes, spraying drying obtains Lactobacillus casei microcapsule bacterium powder afterwards; Wherein, feeding temperature is 25 ℃, 130 ℃ of inlet temperature, and spraying gun pressure is 150kPa.
Wherein, the weight ratio of skim-milk and beta-cyclodextrin is 7: 3.The weight ratio of skim-milk and gelatin is 7: 3.The weight ratio of skim-milk and gum arabic is 6: 4.The weight ratio of the bacterial sediment thing after the lactobacterium casei of wall material and collection is centrifugal is 5: 1.
The present invention has following technique effect:
Method of the present invention improves the resist drying ability of lactobacterium casei by the combination of adopting the various active resist technology, improve the viable count of product, wherein, adopt the pre-lactobacterium casei of cultivating the metabolism regulating measure can repairing damage, improve the viable count before the spraying drying; Adopt heat shock can impel lactobacterium casei to produce some heat shock protein and improve its resist drying ability; Adopt through the composite protectant of screening and optimization and can protect high temperature and dehydration in the lactobacterium casei tolerance spray-drying process; The lactobacterium casei embedding can be reduced in thermal damage and dehydration damage in the spray-drying process by wall material and the microcapsule preparation technology who selects to suit.The survival rate that adopts the lactobacterium casei bacterium powder that method of the present invention makes is all more than 60%, and viable count is all 10
10Individual/more than the g.
Method of the present invention is simple to operate, continuous, quick, can accomplish scale production, and energy consumption is low.Products obtained therefrom is long storage period, packaged convenience.
Embodiment
The present invention is described in detail below in conjunction with specific embodiment.
Embodiment 1
(1) high-density culture: the lactobacterium casei yeast culture thing 10ml aseptic technique after will activating is seeded to 10 liters of MRS liquid nutrient mediums, cultivates 24 hours down at 37 ℃, and it is cultivated viable count and reaches 1.04 * 10
12/ ml.
(2) the lactobacterium casei viable bacteria culture that step (1) high-density culture is obtained cooled off 20 minutes in 4 ℃ of water-baths, centrifugal collection lactobacterium casei bacterial sediment thing, and the bacterial sediment thing weight after the lactobacterium casei that obtains is centrifugal is 68g.
(3) preparation protective material: 0.0136g Sodium Tetraborate, 0.68g gsh are mixed, add the 68ml water dissolution, add 4.08g trehalose, 2.72g sucrose and 0.0272g vitamins C at last, mix.Sterilized 15 minutes, and made the protective material annex solution for 121 ℃.
(4) the protective material annex solution that step (3) is obtained joins in lactobacterium casei that step (2) the collects bacterial sediment thing after centrifugal by aseptic technique; shake to make in 5 minutes and suspend evenly; 4 ℃ leave standstill 1h; afterwards; continue down to cultivate 2 hours in 37 ℃, obtain containing protectant lactobacterium casei thallus suspension liquid.
(5) take by weighing skim-milk 238g, beta-cyclodextrin 102g adds the 1020ml water dissolution.The 8000r/min homogeneous is 5 minutes afterwards, sterilizes 15 minutes, and is cooled to 40 ℃ for 121 ℃.What add that step (4) obtains contains protectant lactobacterium casei thallus suspension liquid and 1.36g Tween 80, and the 5000r/min homogeneous is 10 minutes afterwards.Afterwards, the small-sized spray drier (Japanese physics and chemistry EYELA) of employing 10L carries out spraying drying and obtains Lactobacillus casei microcapsule bacterium powder, wherein, spray-dired input concentration is 25%, and feed rate is 20ml/min, and feeding temperature is 25 ℃, 130 ℃ of inlet temperature, spraying gun pressure 150kPa.
The survival rate of the Lactobacillus casei microcapsule bacterium powder that the employing aforesaid method makes is 67.2%, and viable count is 1.84 * 10
10Individual/g.
Embodiment 2
(1) the lactobacterium casei yeast culture thing 10ml aseptic technique after the activation is seeded to 10 liters of MRS liquid nutrient mediums, cultivates 24 hours down at 37 ℃, and it is cultivated viable count and reaches 1.26 * 10
12/ ml.
(2) the lactobacterium casei viable bacteria culture that step (1) high-density culture is obtained heats 45min in the water-bath of 50 ℃ of heat-shock temperatures, cultivates 1h down at 37 ℃ then.Centrifugal collection lactobacterium casei bacterial sediment thing, obtaining the bacterial sediment thing weight of lactobacterium casei after centrifugal is 71g.
(3) preparation protective material: Sodium Tetraborate 0.0142g, Sodium Glutamate 0.568g, N.F,USP MANNITOL 1.42g and soybean lecithin 3.55g are mixed, add the 71ml water dissolution, add trehalose 0.568g, sucrose 2.84g and 0.0284g vitamins C at last, mix.Sterilized 15 minutes for 121 ℃, it is standby to make the protective material annex solution.
(4) the protective material annex solution that step (3) is obtained joins in lactobacterium casei that step (2) the collects bacterial sediment thing after centrifugal by aseptic technique; shake to make in 5 minutes and suspend evenly; 4 ℃ leave standstill 1h; afterwards; continue down to cultivate 2 hours in 37 ℃, obtain containing protectant lactobacterium casei thallus suspension liquid.
(5) take by weighing skim-milk 248.5g, gelatin 106.5g adds the 1065ml water dissolution.The 8000r/min homogeneous is 5 minutes afterwards.Sterilized 15 minutes, and be cooled to 40 ℃ for 121 ℃.What add that step (4) obtains contains protectant lactobacterium casei thallus suspension liquid and 1.42g Tween 80, and the 5000r/min homogeneous is 10 minutes afterwards.Adopt the small-sized spray drier (Japanese physics and chemistry EYELA) of 10L afterwards.Carry out spraying drying and obtain Lactobacillus casei microcapsule bacterium powder, wherein, input concentration is 25%, and feed rate 20ml/min, feeding temperature are 25 ℃, 130 ℃ of inlet temperature, spraying gun pressure 150kPa.
The survival rate of the lactobacterium casei bacterium powder that the employing aforesaid method makes is 69.8%, and viable count is 1.94 * 10
10Individual/g.
Embodiment 3
(1) the lactobacterium casei yeast culture thing 10ml aseptic technique after the activation is seeded to 10 liters of MRS liquid nutrient mediums, cultivates 24 hours down at 37 ℃, and it is cultivated viable count and reaches 1.35 * 10
12Individual/ml.
(2) the lactobacterium casei viable bacteria culture that step (1) high-density culture is obtained heats 45min in the water-bath of 50 ℃ of heat-shock temperatures, cultivates 1h down at 37 ℃ then.Centrifugal collection lactobacterium casei bacterial sediment thing, obtaining the bacterial sediment thing weight of lactobacterium casei after centrifugal is 75g.
(3) preparation protective material: Sodium Tetraborate 0.0075g, gsh 1.2g, soybean lecithin 3.0g are mixed, add the 75ml water dissolution, add trehalose 0.9g, sucrose 1.5g and 0.015g vitamins C at last, mix.Sterilized 15 minutes for 121 ℃, it is standby to make the protective material annex solution.
(4) the protective material annex solution that step (3) is obtained joins in lactobacterium casei that step (2) the collects bacterial sediment thing after centrifugal by aseptic technique; shake to make in 5 minutes and suspend evenly; 4 ℃ leave standstill 1h; afterwards; continue down to cultivate 2 hours in 37 ℃, obtain containing protectant lactobacterium casei thallus suspension liquid.
(5) take by weighing skim-milk 225g, gum arabic 150g adds the 1125ml water dissolution.The 8000r/min homogeneous is 5 minutes afterwards.Sterilized 15 minutes, and be cooled to 40 ℃ for 121 ℃.What add that step (4) obtains contains protectant lactobacterium casei thallus suspension liquid and 1.50g Tween 80, and the 5000r/min homogeneous is 10 minutes afterwards.Adopt the small-sized spray drier (Japanese physics and chemistry EYELA) of 10L afterwards.Carry out spraying drying and obtain Lactobacillus casei microcapsule bacterium powder, wherein, input concentration is 25%, and feed rate 20ml/min, feeding temperature are 25 ℃, 130 ℃ of inlet temperature, spraying gun pressure 150kPa.
The survival rate of the lactobacterium casei bacterium powder that the employing aforesaid method makes is 72.7%, and viable count is 1.99 * 10
10Individual/g.
Claims (6)
1. a method of utilizing spray drying method for preparation Lactobacillus casei microcapsule bacterium powder is characterized in that, comprises the steps:
(1) high-concentration culturing: the lactobacterium casei yeast culture thing after will activating is that 1% inoculum size is seeded to the MRS liquid nutrient medium with volume fraction, cultivates 24 hours down at 37 ℃, makes the maximum viable count of cultivating reach 10
10Individual/more than the ml;
(2) adopt pre-cultivation or heat shock to handle as the metabolism regulating measure:
Described pre-cultural method is:
The lactobacterium casei viable bacteria culture that step (1) high-density culture is obtained cooled off 10-20 minute in 4 ℃ of water-baths, centrifugal collection lactobacterium casei bacterial sediment thing, afterwards the protective material annex solution is joined in the lactobacterium casei of the collection bacterial sediment thing after centrifugal by aseptic technique, shake to make in 5 minutes and suspend evenly, 4 ℃ leave standstill 1h, afterwards, continue down to cultivate 2 hours, obtain containing protectant lactobacterium casei thallus suspension liquid in 37 ℃;
Described heat shock treatment process is:
The lactobacterium casei viable bacteria culture that step (1) high-density culture is obtained heats 45min in heat-shock temperature is 50 ℃ water-bath, afterwards, cultivate 1h, centrifugal collection lactobacterium casei bacterial sediment things down at 37 ℃; The protective material annex solution is joined in the lactobacterium casei of the collection bacterial sediment thing after centrifugal by aseptic technique, shake to make in 5 minutes and suspend evenly, 4 ℃ leave standstill 1h, obtain containing protectant lactobacterium casei thallus suspension liquid;
Wherein, described protective material is by trehalose, sucrose, Sodium Tetraborate, Sodium Glutamate or gsh, vitamins C is formed, the add-on of the per-cent meter of the bacterial sediment thing weight after the lactobacterium casei that obtains according to collection is centrifugal is: the 0.6-1.2% trehalose, 2-8% sucrose, the 0.01-0.03% Sodium Tetraborate, 0.8-1.5% Sodium Glutamate or 0.8-1.6% gsh, the 0.02-1.0% vitamins C, and obtain: earlier with Sodium Tetraborate by following method, Sodium Glutamate or gsh mix, the water dissolution of the bacterial sediment thing identical weight after the lactobacterium casei that adds and collect is centrifugal, add trehalose at last, sucrose and vitamins C, mix, sterilized 15 minutes for 121 ℃ afterwards, make the protective material annex solution;
(3) preparation Lactobacillus casei microcapsule:
Select for use in the mixture of mixture, skim-milk and gum arabic of mixture, skim-milk and gelatin of skim-milk and beta-cyclodextrin any as the wall material, contain protectant lactobacterium casei thallus suspension liquid as core;
Add the water dissolution of 3 times of wall material weight in the wall material, the 8000r/min homogeneous is 5 minutes afterwards, sterilizes 15 minutes, and is cooled to 40 ℃ for 121 ℃; Add the Tween 80 that contains protectant lactobacterium casei thallus suspension liquid and wall material solution weight 0.1% again, 5000r/min second homogenate 10 minutes, spraying drying obtains Lactobacillus casei microcapsule bacterium powder afterwards; Wherein, spray-dired feeding temperature is 25 ℃, 130 ℃ of inlet temperature, and spraying gun pressure is 150kPa.
2. the method for utilizing spray drying method for preparation Lactobacillus casei microcapsule bacterium powder according to claim 1 is characterized in that the weight ratio of skim-milk and beta-cyclodextrin is 7: 3.
3. the method for utilizing spray drying method for preparation Lactobacillus casei microcapsule bacterium powder according to claim 1 is characterized in that the weight ratio of skim-milk and gelatin is 7: 3.
4. the method for utilizing spray drying method for preparation Lactobacillus casei microcapsule bacterium powder according to claim 1 is characterized in that the weight ratio of skim-milk and gum arabic is 6: 4.
5. the method for utilizing spray drying method for preparation Lactobacillus casei microcapsule bacterium powder according to claim 1 is characterized in that the weight ratio of the bacterial sediment thing after the lactobacterium casei of wall material and collection is centrifugal is 5: 1.
6. the method for utilizing spray drying method for preparation Lactobacillus casei microcapsule bacterium powder according to claim 1; it is characterized in that; also contain N.F,USP MANNITOL and soybean lecithin in the described protective material; the add-on of the per-cent meter of the bacterial sediment thing weight after the lactobacterium casei that obtains according to collection is centrifugal is: 1-4% N.F,USP MANNITOL; the 4-8% soybean lecithin; protective material obtains by following method: earlier with Sodium Tetraborate; Sodium Glutamate or gsh; N.F,USP MANNITOL and soybean lecithin mix; the water dissolution of the bacterial sediment thing identical weight after the lactobacterium casei that adds and collect is centrifugal; add trehalose at last; sucrose and vitamins C; mix; sterilized 15 minutes for 121 ℃ afterwards, make the protective material annex solution.
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