Fermented celery powder and preparation method thereof

Abstract

The invention discloses fermented celery powder and a preparation method thereof, and belongs to the technical field of microbial fermentation engineering. Celery is directly pulped, cellulose is used for pre-enzymolysis, cellulose in celery pulp is moderately decomposed, nitrate wrapped by cell walls of the cellulose is released, and then nitrate is fermented and reduced into nitrite through nitrate reducing bacteria. Through the synergistic effect of enzymes and bacteria, nitrate in the celery pulp is efficiently recycled and converted, and the overall yield of nitrite in the fermented celery powder is gt; and the input-output requirements of industrial production are met. Meanwhile, the fermented celery powder retains more vitamin C and the like, is used for pickling meat products instead of nitrous acid, has remarkable effects of corrosion prevention, bacteriostasis, color stabilization, taste and flavor improvement and nitrite residue reduction, and can be used as a green food additive to completely replace chemically synthesized nitrite.

Description

Technical Field

[0001] This invention relates to the field of microbial fermentation engineering technology, specifically to a fermented celery powder and its preparation method. Background Technology

[0002] With the increasing awareness of food safety, the demand for green, ecological, and healthy food is growing, with the annual growth rate of demand for organic green food exceeding 20%. Since organic green food does not allow the addition of chemically synthesized food additives, the green food additive industry has experienced rapid development in recent years.

[0003] Nitrites are a commonly used chemically synthesized food preservative additive, possessing antibacterial, antioxidant, and flavor-enhancing effects on meat products. Especially during meat processing, adding nitrites can significantly improve the color of meat products, substantially inhibit the growth of foodborne pathogens, and provide long-term preservation. Nitrites play a crucial role in the processing and preservation of meat products. However, according to publicly available reports, chemically synthesized nitrites can be converted into carcinogenic nitrosamines under certain conditions, and long-term excessive intake may increase the risk of cancer. my country's Green Food and Food Additive Usage Guidelines (NY / T 392-2023) also clearly stipulate that the use of chemically synthesized food additives such as nitrates or nitrites is prohibited in the production of green and organic foods. Therefore, developing nitrite substitutes has become an important branch of the development of green food additives.

[0004] Celery is rich in nitrates, generally exceeding 1000 mg / kg, making it a natural biological source of nitrates. The nitrates in celery are converted into nitrites through microbial action, which can be used as a natural green food additive to replace chemically synthesized nitrites. If chemically synthesized nitrites are added directly during meat curing, it generally results in excessive nitrite residue, while using celery powder as an additive results in much lower nitrite residue.

[0005] Currently, celery is being used as a preservative in the curing of meat to meet the requirements for green food additives. One method involves adding celery powder and microbial agents to the meat, allowing the celery powder to gradually ferment and convert nitrates into nitrites. However, this method is difficult to control during fermentation, making it challenging to control the dosage. Another method involves fermenting celery with nitrate-reducing bacteria to produce fermented celery powder containing natural nitrites. This method is much more convenient and allows for precise dosage control. This fermented celery powder, rich in natural nitrites, offers significant advantages as a green, healthy, and safe food additive in replacing chemically synthesized nitrites.

[0006] Current development of fermented celery powder mainly focuses on matching suitable bacterial strains to the pressed celery juice to improve the nitrite conversion rate. For example, Chinese invention patent publication number CN116716205A discloses a method for producing nitrite-rich celery powder, using Staphylococcus camosus (accession number CCTCC NO: M 20221939) as the fermentation strain to ferment celery juice, achieving a nitrite conversion rate higher than 95%. Li Peiran of Tianjin Agricultural College, in his study "Preparation of Fermented Celery Powder and Its Application Effect in Enemas," improved the nitrite conversion rate by screening bacterial strains, and the WBL-45 strain from SACCO Company showed a significant effect in improving nitrite conversion.

[0007] However, there are still certain barriers to the industrial production of fermented celery powder using existing technologies. Celery juicing produces a large amount of celery pulp, and the byproducts of celery pulp contain some nitrates, vitamin C, polyphenols, etc., which cannot be fully extracted and utilized by the juicer, resulting in great waste, reduced overall yield, and serious nutrient loss, which is not conducive to industrial production. Summary of the Invention

[0008] To adapt to industrial production and fully utilize the nitrates, vitamin C, polyphenols, and other components in celery to improve yield, this invention proposes a fermented celery powder and its preparation method. Specifically, the celery is completely crushed into a pulp. Through enzyme-microbe synergy, cellulase is used to moderately decompose the cellulose in the celery pulp, fully releasing effective components such as nitrates, vitamin C, and polyphenols. Further fermentation with microbial inoculum significantly increases the yield of nitrite in the fermented celery powder.

[0009] A method for preparing fermented celery powder mainly includes the following steps:

[0010] S1. Celery pulp preparation: Wash fresh celery, drain, and crush it through a 10-mesh sieve to obtain celery pulp;

[0011] S2. Preparation of celery pulp pre-enzymatic hydrolysate: Celery pulp and water were added to the enzymatic hydrolysis tank at a mass ratio of 1:(0.2-0.4), and then cellulase was added. The cellulase activity of the contents of the enzymatic hydrolysis tank was controlled at 2-5 u / ml and the pH value was 4.5-5.5. The mixture was then sealed and allowed to stand for 10-15 hours at 40-45℃. The mixture was then filtered by pressing using a plate and frame filter press to obtain the celery pulp pre-enzymatic hydrolysate.

[0012] S3. Fermentation broth preparation: Add the pre-enzymatic hydrolysate of celery pulp to the fermenter, then inoculate with nitrate-reducing bacteria at a rate of 1.0–2.0 × 10⁻⁶. 7CFU / mL; control the fermenter temperature to be constant at 30-35℃, adjust the pH value to 6.0-6.5, stir at low speed, control the air inlet and exhaust valves to adjust the oxygen content of the slurry to 8-10mg / L, after fermentation for 8 hours, check the nitrite content every 1 hour, and stop fermentation when the nitrite content rises below 10mg / kg;

[0013] S4. Preparation of fermented celery powder: The fermentation liquid is squeezed out through a plate and frame filter press, 2% activated carbon is added to the filtrate for decolorization, and then concentrated and vacuum dried in a vacuum concentrator to obtain fermented celery powder.

[0014] Preferably, the celery is selected from either Chinese celery or Western celery.

[0015] Preferably, the cellulase activity is 10,000 to 20,000 u / g. The cellulase has the CAS number 9012-54-8, which degrades insoluble cellulose and can disrupt the cell wall of celery, thereby promoting the release of nitrates.

[0016] Preferably, in step S2, citric acid is used to adjust the pH to 4.5–5.5. By adjusting to a weakly acidic environment, cellulase activity is released. To prevent over-enzymatic hydrolysis, the pH should not be too low.

[0017] Preferably, the sealed static aging in step S2 is controlled to be 12 hours. The purpose of aging is to utilize cellulase to moderately hydrolyze the cellulose, so as to facilitate the full release of the nitrates encapsulated within, thereby increasing the yield of nitrites and meeting the technical requirements for input and output in industrial production. Excessive aging time will lead to over-enzymatic hydrolysis of the celery pulp, increasing sugar content and affecting subsequent fermentation. Furthermore, the quality of the fermented celery powder obtained from excessive hydrolysis will decrease.

[0018] Preferably, the pressure range of the plate and frame filter press in step S2 is 0.2 to 0.4 MPa, and the pore size of the filter cloth of the plate and frame filter press is 100 µm.

[0019] Preferably, the nitrate-reducing bacteria mentioned in step S3 are Staphylococcus carnosus (CCTCC NO: M 20221939) strains inoculated into the culture medium and cultured at 37°C for 18 hours to obtain a stable viable count of 10. 10 High-density nitrate-reducing bacteria at the CFU / mL level; specifically, the culture medium contains the following nutrients: glucose 2 g / L, soybean peptone 18 g / L, beef extract 22 g / L, and sodium chloride 2 g / L. The high-density nitrate-reducing bacteria can be directly added to the solution, meeting the needs of industrial production.

[0020] Preferably, in step S3, ammonia is used to adjust the fermentation pH to 6.5. The pH should not be too low, as a low pH will cause the generated nitrite to undergo acid degradation, affecting the final nitrite content.

[0021] Preferably, the pressure range of the plate and frame filter press in step S3 is 0.2 to 0.4 MPa, and the pore size of the filter cloth in the plate and frame filter press is 20 µm.

[0022] Preferably, the vacuum decompression concentrator and vacuum dryer rapidly evaporate moisture at 40-60°C, thereby avoiding heat loss of vitamin C and other nutrients in the fermented celery powder.

[0023] This invention also protects a fermented celery powder prepared by the above method. The nitrate content in celery is mainly concentrated in the stems and leaves. The cellulose cell walls of the stems and leaves easily trap and fix nitrates, preventing their full release and conversion. This invention directly pulps celery, utilizes cellulase for pre-enzymatic hydrolysis to moderately decompose the cellulose in the celery pulp, releasing the nitrates trapped by the cellulose cell walls. Then, nitrate-reducing bacteria ferment the nitrates to reduce them to nitrites. Through the synergistic effect of enzymes and bacteria, efficient conversion of nitrates in the celery pulp is achieved, increasing the yield of nitrites in the fermented celery powder. Simultaneously, vitamin C, polyphenols, and flavonoids in celery are fully extracted, giving the fermented celery powder rich nutritional value. When the fermented celery powder is used as an additive in cured meats, vitamin C and other nutrients can moderately block the conversion of nitrites to the carcinogen nitrosamine, reducing nitrites to nitric oxide. This not only improves the color stability of cured meat products but also enhances their taste and flavor, while reducing nitrite residue. This ensures the comprehensive advantages of fermented celery powder in replacing chemically synthesized nitrites in terms of preservation, color protection, and improved taste.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] (1) This invention does not use celery juice, but instead crushes all the celery for processing, so that more nitrates, vitamin C and polyphenols can be utilized, thereby improving the overall yield of production and avoiding the waste of raw materials in industrial production.

[0026] (2) This invention utilizes the synergistic effect of enzymes and bacteria to moderately enzymatically hydrolyze cellulose, fully releasing effective components such as nitrates, vitamin C, and polyphenols, thereby making full use of nitrates in celery, significantly improving the fermentation conversion efficiency and yield of nitrites, and reducing excessive dependence on strains.

[0027] (3) The fermented celery powder of the present invention retains more vitamin C and polyphenols, and can replace nitrite in the curing of meat products. It has significant effects on preservation and antibacterial properties, color stabilization, taste and flavor enhancement, and nitrite residue reduction. It can be used as a green food additive to completely replace nitrite. Detailed Implementation

[0028] The specific implementation methods of this invention are described in detail below. These embodiments are merely some, not all, of the embodiments of this invention. All other embodiments obtained by those skilled in the art based on the technical concepts of the embodiments of this invention without creative effort are within the scope of protection of this invention.

[0029] Unless otherwise stated, the equipment, testing instruments, and testing methods involved in the following implementation processes are all common knowledge understood by those skilled in the art; the relevant testing standards referred to are GB 5009.33-2016 "Determination of Nitrite and Nitrate in Food", GB 5009.228-2016 "Determination of Volatile Basic Nitrogen in Food", and GB 5009.86-2016 "Determination of Ascorbic Acid in Food".

[0030] All equipment involved was sterilized at 120℃ for 30 minutes before use.

[0031] When calculating yields, nitrates and nitrites are measured as sodium nitrate and sodium nitrite, respectively.

[0032] Raw material preparation involved:

[0033] Cellulase: Enzyme activity 10,000 u / g, produced by Nanning Pangbo Biotechnology Co., Ltd.

[0034] Celery: average nitrate content 1240 mg / kg.

[0035] Staphylococcus carnosus: accession number CCTCC NO:M 20221939, published by Chinese invention patent CN116716205A on September 8, 2023, applicants are Zhejiang Xinyinxiang Biotechnology Co., Ltd. and Zhejiang Yitao Food Technology Co., Ltd.

[0036] Preparation of nitrate-reducing bacteria: Staphylococcus carnosus (CCTCC NO: M 20221939) was inoculated into a culture medium with the following nutrients: glucose 2 g / L, soybean peptone 18 g / L, beef extract 22 g / L, and sodium chloride 2 g / L. The medium was incubated at 37°C for 18 hours until the viable bacterial count stabilized at 10⁻⁶. 10 This high-density nitrate-reducing bacteria, at the CFU / mL level, can be used directly.

[0037] Example 1

[0038] S1. Celery pulp preparation: Wash, drain, and crush 100kg of fresh celery to pass through a 10-mesh sieve to obtain celery pulp;

[0039] S2. Preparation of pre-enzymatic hydrolysate of celery pulp: Celery pulp and water were added to an enzymatic hydrolysis tank at a mass ratio of 1:0.3, and then cellulase was added. The cellulase activity of the contents of the enzymatic hydrolysis tank was controlled at 3u / ml. The pH value was adjusted to 5.0 with citric acid. The mixture was then allowed to stand in a sealed container at 45℃ for 12 hours. The mixture was then filtered by pressing using a plate and frame filter press with a pressure of 0.3MPa and a filter cloth pore size of 100µm to obtain the pre-enzymatic hydrolysate of celery pulp.

[0040] S3. Fermentation broth preparation: Add the celery pulp pre-hydrolysate to the fermentation tank, then inoculate with 10... 10 High-density nitrate-reducing bacteria at the CFU / mL level, with an inoculation amount of 1.25 × 10⁻⁶. 7 CFU / mL; The fermentation tank temperature was kept constant at 35℃, the pH was adjusted to 6.5 using ammonia, and the mixture was stirred at a low speed of 50 rpm. The oxygen content of the slurry was adjusted to 10 mg / L by controlling the air intake and exhaust valves. After 8 hours of fermentation, the nitrite content was tested every 1 hour, as shown in Table 1. When the nitrite content increased by less than 10 mg / kg after 10 hours of fermentation, the fermentation was stopped.

[0041] S4. Preparation of fermented celery powder: The fermentation liquid was squeezed out through a plate and frame filter press. The plate and frame filter press was pressurized at 0.3 MPa and the filter cloth pore size was 20 µm. 2% activated carbon was added to the filtrate for decolorization. The filtrate was concentrated in a vacuum concentrator and vacuum dried at -0.08 MPa. The filtrate was dried at 50℃ to obtain 7.82 kg of fermented celery powder.

[0042] Example 2

[0043] S1. Celery pulp preparation: Wash, drain, and crush 100kg of fresh celery to pass through a 10-mesh sieve to obtain celery pulp;

[0044] S2. Preparation of pre-enzymatic hydrolysate of celery pulp: Celery pulp and water were added to an enzymatic hydrolysis tank at a mass ratio of 1:0.4, and then cellulase was added. The cellulase activity of the contents of the enzymatic hydrolysis tank was controlled at 5 u / ml. The pH value was adjusted to 4.5 with citric acid. The mixture was then allowed to stand in a sealed container at 40℃ for 15 h. The mixture was then filtered by pressing using a plate and frame filter press with a pressure of 0.3 MPa and a filter cloth pore size of 100 µm to obtain the pre-enzymatic hydrolysate of celery pulp.

[0045] S3. Fermentation broth preparation: Add the celery pulp pre-hydrolysate to the fermentation tank, then inoculate with 10... 10 High-density nitrate-reducing bacteria at the CFU / mL level, with an inoculation amount of 1.86 × 10⁻⁶. 7FU / mL; control the fermenter temperature to be constant at 35℃, adjust the pH value to 6.0 using ammonia water, stir at a low speed of 50rpm, and control the air inlet and outlet valves to adjust the oxygen content of the slurry to 8mg / L. After 8 hours of fermentation, test the nitrite content every 1 hour, as shown in Table 1; after 10 hours of fermentation, if the increase in nitrite content is less than 10mg / kg, stop fermentation.

[0046] S4. Preparation of fermented celery powder: The fermentation liquid was squeezed out through a plate and frame filter press. The plate and frame filter press was pressurized at 0.3 MPa and the filter cloth pore size was 20 µm. 2% activated carbon was added to the filtrate for decolorization. The filtrate was concentrated in a vacuum concentrator and vacuum dried at -0.08 MPa. The filtrate was dried at 50℃ to obtain 7.91 kg of fermented celery powder.

[0047] Comparative Example 1

[0048] This scheme follows the basic process of the embodiment, but does not perform cellulase enzymatic hydrolysis pretreatment on the celery pulp. The specific implementation method is as follows:

[0049] S1. Celery pulp preparation: Wash, drain, and crush 100kg of fresh celery to pass through a 10-mesh sieve. Mix the celery pulp and water at a mass ratio of 1:0.3. Use a plate and frame filter press to pressurize and filter the mixture. The plate and frame filter press is pressurized at 0.3MPa and the filter cloth has a pore size of 100µm to obtain celery juice.

[0050] S3. Fermentation broth preparation: Add celery juice to the fermentation tank, then inoculate with 10... 10 High-density nitrate-reducing bacteria at the CFU / mL level, with an inoculation amount of 1.25 × 10⁻⁶. 7 CFU / mL; control the fermenter temperature to be constant at 35℃, adjust the pH value to 6.5, stir at a low speed of 50rpm, control the air inlet and outlet valve to adjust the oxygen content of the slurry to 10mg / L, after fermentation for 8 hours, test the nitrite content every 1 hour as shown in Table 1; when the nitrite content rises below 10mg / kg after 12 hours of fermentation, stop fermentation.

[0051] S4. Preparation of fermented celery powder: The fermentation liquid was squeezed out through a plate and frame filter press. The plate and frame filter press was pressurized at 0.3 MPa and the filter cloth pore size was 20 µm. 2% activated carbon was added to the filtrate for decolorization. The filtrate was concentrated in a vacuum concentrator and vacuum dried at -0.08 MPa. The filtrate was dried at 50℃ to obtain 6.46 kg of fermented celery powder.

[0052] Table 1: Changes in nitrite content during fermentation

[0053]

[0054] By comparing the fermentation processes of Examples 1-2 and Comparative Example 1 in Table 1, it is found that the present invention, after celery is pulped, is hydrolyzed by cellulase before fermentation, which shortens the fermentation time and increases the fermentation efficiency, and can complete the fermentation in 9 hours. In contrast, celery pulp without pre-hydrolysis ferments more slowly and has a lower nitrite content.

[0055] Effect Experiment

[0056] I. Yield assessment of nitrite in fermented celery powder:

[0057] Each implementation plan used 100 kg of celery raw material, which theoretically contained 124 g of nitrate and could theoretically be converted into 101 g of nitrite. The yield of nitrite in fermented celery powder was calculated by detecting the mass M (kg) of fermented celery powder and the nitrite content λ (g / kg) of fermented celery powder. The yield (Y) was calculated as (M×λ) / 101×100%. The results are shown in Table 2.

[0058] The vitamin C content of fermented celery powder was determined according to GB 5009.86-2016 "Determination of ascorbic acid in food". The results are shown in Table 2.

[0059] Table 2: Yield of Nitrite from Fermented Celery Powder

[0060]

[0061] Table 2 shows the yield of nitrite in fermented celery powder. This invention, through pretreatment of celery pulp with cellulase, effectively breaks down nitrates, vitamin C, and other substances fixed in the cellulosic cell walls, allowing them to be fully released. Based on nitrite in fermented celery powder, the overall yield is increased from the conventional 74.71% to over 87%, and the vitamin C content is also high. This is extremely important for reducing losses and promoting the industrialization of fermented celery powder.

[0062] II. Evaluation of the effectiveness of fermented celery powder in replacing nitrites for meat curing:

[0063] According to GB 2760-2024 "National Food Safety Standard for the Use of Food Additives", the maximum permitted amount of nitrite (calculated as sodium nitrite) in cured meat is 0.15 g / kg, and the maximum residue limit is 30 mg / kg. The fermented celery powder obtained in each embodiment of this invention is used to equivalently replace sodium nitrite in sausage curing, with an equivalent addition of 0.10 g / kg of sodium nitrite as the standard. Pork (lean to fat ratio 7:3) is minced using a meat grinder, sterilized with daqu liquor for 30 minutes, then mixed with fermented celery powder (nitrite), sugar, salt, Sichuan pepper powder, and other auxiliary materials, and then packaged to complete the sausage curing process. Sausages cured with and without sodium nitrite are used as control samples. The weight distribution of each curing auxiliary material based on pork is shown in Table 3.

[0064] Table 3: Marinating Ingredients

[0065]

[0066] Table 3 shows the changes in residual nitrite and volatile basic nitrogen (TVB-N) content (automatic Kjeldahl method) during the 15-day curing process of sausages cured with different ingredients. Generally, the TVB-N content increases with prolonged curing time. Cured meat undergoes putrefaction due to bacterial action, and protein decomposition produces nitrogenous substances such as ammonia and amines, which become volatile basic nitrogen. Lower TVB levels indirectly indicate better preservative effect. The test results are shown in Table 4.

[0067] Table 4: Nitrite Residue and Volatile Basic Nitrogen Content

[0068]

[0069] Table 4 shows the detection of nitrite residue and TVB-N in cured sausages. Compared with chemically synthesized sodium nitrite, the fermented celery powder of this invention has lower nitrite residue and lower TVB-N value, and significantly inhibits protein putrefaction and decomposition. It can completely replace sodium nitrite as a preservative and color-preserving additive in cured meat products.

Claims

1. A method for preparing fermented celery powder, characterized in that, Includes the following steps: S1. Celery pulp preparation: Wash fresh celery, drain, and crush it through a 10-mesh sieve to obtain celery pulp; S2. Preparation of celery pulp pre-enzymatic hydrolysate: Celery pulp and water were added to the enzymatic hydrolysis tank at a mass ratio of 1:(0.2-0.4), and then cellulase was added. The cellulase activity of the contents of the enzymatic hydrolysis tank was controlled at 2-5 u / ml and the pH value was 4.5-5.

5. The mixture was then sealed and allowed to stand for 10-15 hours at 40-45℃. The mixture was then filtered by pressing using a plate and frame filter press to obtain the celery pulp pre-enzymatic hydrolysate. S3. Fermentation broth preparation: Add the pre-enzymatic hydrolysate of celery pulp to the fermenter, then inoculate with nitrate-reducing bacteria at a rate of 1.0–2.0 × 10⁻⁶. 7 CFU / mL; control the fermenter temperature to be constant at 30-35℃, adjust the pH value to 6.0-6.5, stir at low speed, control the air inlet and exhaust valves to adjust the oxygen content of the slurry to 8-10mg / L, after fermentation for 8 hours, check the nitrite content every 1 hour, and stop fermentation when the nitrite content rises below 10mg / kg; S4. Preparation of fermented celery powder: The fermentation liquid is squeezed out through a plate and frame filter press, 2% activated carbon is added to the filtrate for decolorization, and then concentrated and vacuum dried in a vacuum concentrator to obtain fermented celery powder.

2. The method for preparing fermented celery powder according to claim 1, characterized in that, The celery used is either Chinese celery or Western celery.

3. The method for preparing fermented celery powder according to claim 1, characterized in that, The activity of the cellulase described in step S2 is 10,000 to 20,000 u / g.

4. The method for preparing fermented celery powder according to claim 1, characterized in that, In step S2, citric acid is used to adjust the pH value to 4.5–5.

5.

5. The method for preparing fermented celery powder according to claim 1, characterized in that, The sealed static storage in step S2 is controlled for 12 hours.

6. The method for preparing fermented celery powder according to claim 1, characterized in that, The pressure range of the plate and frame filter press in step S2 is 0.2 to 0.4 MPa, and the pore size of the filter cloth in the plate and frame filter press is 100 µm.

7. The method for preparing fermented celery powder according to claim 1, characterized in that, The nitrate-reducing bacteria mentioned in step S3 are Staphylococcus carnosus inoculated into the culture medium and cultured at 37°C for 18 hours, resulting in a stable viable count of 10⁻⁶. 10 High-density nitrate-reducing bacteria at CFU / mL level; the preservation number of the Staphylococcus carnosus is CCTCC NO:M 20221939; the nutrients in the culture medium are: glucose 2g / L, soybean peptone 18g / L, beef extract powder 22g / L, and sodium chloride 2g / L.

8. The method for preparing fermented celery powder according to claim 1, characterized in that, In step S3, ammonia water is used to adjust the fermentation pH to 6.

5.

9. The method for preparing fermented celery powder according to claim 1, characterized in that, The pressure range of the plate and frame filter press in step S3 is 0.2 to 0.4 MPa, and the pore size of the filter cloth in the plate and frame filter press is 20 µm.

10. Fermented celery powder obtained by the preparation method according to any one of claims 1-9.

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