Method for extracting fresh moringa oleifera leaf protein by adding molasses for fermentation

Abstract

The invention provides a method for extracting protein from fresh moringa oleifera leaves by adding molasses for fermentation. The method comprises the following steps: S1, pretreating fresh moringa oleifera leaves; s2, adding water and pulping; s3, filtering with a nylon net filter bag, and separating filter residues from filtrate; cleaning the beater, mixing a cleaning solution with filter residues, filtering through a nylon net filter bag, repeating for 3 times, and collecting all filtrate; s4, adding molasses into the collected filtrate, and uniformly mixing; performing anaerobic fermentation in a constant-temperature incubator at 25-45 DEG C for 35-45 hours to obtain fermentation liquor; s5, placing the fermentation liquor in a centrifugal machine, centrifugally separating leaf protein precipitate and supernate, and pouring out the supernate to obtain leaf protein precipitate; and S6, freeze-drying the leaf protein precipitate in a freeze dryer at-80 DEG C for 12 hours to obtain a leaf protein finished product. The extraction rate, the leaf protein yield and the crude protein content of the horseradish tree leaf protein extracted by the method are higher and are superior to those of other extraction methods.

Description

Technical Field

[0001] The invention belongs to the technical field of deep processing and high-value preparation of moringa leaves, and specifically relates to a method for extracting fresh moringa leaf protein by adding molasses and fermenting. Background Art

[0002] Leaf protein refers to a green protein concentrate with certain functionality extracted from fresh green plant leaves through processes such as wall breaking, juicing, and separation. According to different precipitation processes, the crude leaf protein extracted is divided into albumin and green protein. Among them, the main component of albumin is soluble protein, which is used as a human food additive; the main component of green protein is insoluble protein, which is generally used as livestock feed.

[0003] Moringa: Moringa family, Moringales, native to India, also known as drumstick tree, miracle tree, traditional moringa grows in eastern Africa and northern India. Moringa seeds and leaves are rich in nutrients. It is a perennial tropical deciduous tree with about 14 species in the world. The three most commonly eaten species are the following: Indian traditional moringa (Moringa oleifera Lam.), Indian improved moringa (improved species from TN Agricultural University, India, early-maturing and with high pod yield) and African moringa (originally only grown near Lake Turkana in Kenya and southwestern Ethiopia). Moringa leaf is a high-calcium plant product, with calcium content more than 4 times that of milk. In addition, it also contains high trace elements such as potassium, iron, and zinc, and contains protein and amino acids similar to soybeans. It is a functional food containing complete nutrients. A person only needs to eat about 5 grams a day to supply the body's daily nutritional needs. Therefore, it can supplement calcium, iron, potassium, and zinc, and can provide sufficient nutritional needs for weight loss consumers without damaging the body due to dieting. According to measurements, the calcium contained in Moringa dry leaf powder is 4 times that of milk, protein is 2 times that of milk, potassium is 3 times that of bananas, iron is 3 times that of spinach, vitamin C is 7 times that of citrus, and vitamin A (B-carotene) is 4 times that of carrots. Just 3 tablespoons of Moringa leaf powder contain 270% of the daily vitamin A, 42% of protein, 125% of calcium, 70% of iron, and 22% of vitamin C required by children.

[0004] The development and extraction of leaf protein can be traced back to the 1770s internationally. Foreign experts used alcohol to extract colored flocculants (leaf protein) from green plants. After that, people began to study the preparation of leaf protein, improve the process, and establish a leaf protein extraction factory. After the 1990s, the scope of leaf protein research further developed from simple leaf protein crude products to high-end leaf protein refined products, food, medical and health products and fine chemical products, showing the characteristics of all-round, multi-level and comprehensive research and development. my country has rich leaf protein resources, but the research on leaf protein started late. Since the late 1980s, it has begun to carry out comprehensive research on resources, production technology application, efficiency, etc., and achieved corresponding results. There is no essential difference between woody plant leaf protein and herbaceous plant leaf protein in composition and extraction technology. At present, acid-base heating method, salting-out method, ultrafiltration method, etc. are mostly used, but they all have shortcomings. Summary of the invention

[0005] The invention aims to provide a method for extracting protein from fresh Moringa leaves by adding molasses and fermenting. Lactic acid bacteria in the fresh Moringa leaves are enriched by adding molasses and fermenting, so that the pH value is reduced to reach the isoelectric point during the fermentation process, the protein is precipitated, and the extraction rate of leaf protein is improved.

[0006] In order to achieve the above object, the present invention provides the following technical solutions:

[0007] A method for extracting fresh Moringa leaf protein by adding molasses and fermenting, comprising the following steps:

[0008] S1: Pre-treatment of fresh Moringa leaves;

[0009] S2: placing the pre-treated fresh Moringa leaves into a pulping machine, adding water to pulp, and obtaining freshly squeezed leaf pulp;

[0010] S3: filtering the freshly squeezed leaf pulp in step S2 through a nylon mesh filter bag to separate the filter residue and the filtrate; cleaning the pulper, mixing the cleaning liquid with the filter residue and filtering through a nylon mesh filter bag, repeating this process three times, and collecting all the filtrate;

[0011] S4: adding molasses to the filtrate collected in step S3, mixing evenly to obtain a mixed solution; then placing the mixed solution in a constant temperature incubator at 25-45° C. for anaerobic fermentation for 35-45 hours to obtain a fermentation liquid;

[0012] S5: placing the fermentation liquid in step S4 in a centrifuge, centrifuging to separate the leaf protein precipitate and the supernatant, and discarding the supernatant to obtain the leaf protein precipitate;

[0013] S6: The leaf protein precipitate obtained in step S5 is placed in a -80°C freeze dryer and freeze-dried for 12 hours to obtain a leaf protein product.

[0014] Preferably, the pretreatment process of step S1 is: washing the picked fresh Moringa leaves with clean water to remove impurities such as rotten leaves and sand; then cutting the washed fresh Moringa leaves into small segments of 1-2 cm.

[0015] Preferably, in step S2, the solid-liquid ratio of fresh Moringa leaves to water is 1:12-15; and the pulping is performed at a rate of 500 r / min for 2-3 min at 25°C.

[0016] Preferably, the pore size of the nylon mesh in step S3 is 300 mesh.

[0017] Preferably, the molasses in step S4 is sugarcane molasses; the molasses accounts for 5-10% of the mass of the filtrate; and the mixture is manually stirred at 25° C. for 3 minutes until uniform.

[0018] Preferably, in step S5, centrifugation is performed at 4500-6000 r / min for 10-20 min.

[0019] Preferably, in step S6, the leaf protein precipitate is placed in a -60°C freeze dryer and freeze-dried for 12 hours.

[0020] Compared with the prior art, the advantages and beneficial effects of the present invention are:

[0021] The present invention directly uses new Moringa leaves as raw materials to extract and process leaf protein, and compared with the previous research using Moringa leaf powder as the test material, the actual production cost is lower. By pre-treating fresh Moringa leaves, beating and collecting the filtrate, adding molasses and anaerobic fermentation in a constant temperature box to enrich the lactic acid bacteria of the fresh Moringa leaves themselves, so that the pH is reduced to the isoelectric point during the fermentation process, so that the protein is precipitated. The processing process is safe and simple, without chemical pollution, pure and natural, and no waste is generated, and all by-products can be recycled for secondary use. The method extracts Moringa leaf protein with an extraction rate of up to 59.7%, a leaf protein yield of up to 6.9%, a crude protein content of up to 43.5%, complete extraction and a high extraction rate of leaf protein concentrate, and a high protein content, which can serve as a high-quality protein feed. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 : Response surface diagram and contour diagram of the interaction between the three factors in Example 3 on the leaf protein extraction rate. DETAILED DESCRIPTION

[0023] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0024] The present embodiment provides a method for extracting fresh Moringa leaf protein by adding molasses and fermenting, comprising the following steps:

[0025] S1: Wash the freshly picked Moringa leaves with clean water to remove impurities such as rotten leaves and sand; then cut the washed Moringa leaves into small pieces of 1-2 cm;

[0026] S2: placing the pretreated fresh Moringa leaves into a pulper, adding water according to a solid-liquid ratio of 1:12 for pulping, and pulping at a rate of 500 r / min for 2 min at 25° C. to obtain freshly squeezed leaf pulp;

[0027] S3: filtering the freshly squeezed leaf pulp in step S2 through a 280-mesh nylon mesh filter bag to separate the filter residue and the filtrate; cleaning the pulper, mixing the cleaning liquid and the filter residue and filtering again through a 280-mesh nylon mesh filter bag, repeating twice, and collecting all the filtrate;

[0028] S4: adding molasses accounting for 5% of the mass of the filtrate to the filtrate collected in step S3; stirring for 3 minutes at 25°C until uniform to obtain a mixed solution; and then placing in a constant temperature incubator at 25°C for anaerobic fermentation for 35 hours to obtain a fermentation liquid;

[0029] S5: placing the fermentation liquid in step S4 in a centrifuge, centrifuging at 4500 r / min for 10 min to separate the leaf protein precipitate and the supernatant, and discarding the supernatant to obtain the leaf protein precipitate;

[0030] S6: The leaf protein precipitate obtained in step S5 is placed in a -60°C freeze dryer and freeze-dried for 12 hours to obtain a leaf protein product.

[0031] Example 2

[0032] The present embodiment provides a method for extracting fresh Moringa leaf protein by adding molasses and fermenting, comprising the following steps:

[0033] S1: Wash the freshly picked Moringa leaves with clean water to remove impurities such as rotten leaves and sand; then cut the washed Moringa leaves into small pieces of 1-2 cm;

[0034] S2: placing the pretreated fresh Moringa leaves into a pulper, adding water according to a solid-liquid ratio of 1:15 for pulping, and pulping at a rate of 500 r / min for 3 min at 25° C. to obtain freshly squeezed leaf pulp;

[0035] S3: filtering the freshly squeezed leaf pulp in step S2 through a 320-mesh nylon mesh filter bag to separate the filter residue and the filtrate; cleaning the pulper, mixing the cleaning liquid and the filter residue and filtering again through a 320-mesh nylon mesh filter bag, repeating 5 times, and collecting all the filtrate;

[0036] S4: adding molasses accounting for 10% of the mass of the filtrate to the filtrate collected in step S3; stirring for 3 minutes at 25°C until the mixture is uniform to obtain a mixed solution; and then placing the mixture in a constant temperature incubator at 45°C for anaerobic fermentation for 45 hours to obtain a fermentation liquid;

[0037] S5: placing the fermentation liquid in step S4 in a centrifuge, centrifuging at 6000 r / min for 20 min to separate the leaf protein precipitate and the supernatant, and discarding the supernatant to obtain the leaf protein precipitate;

[0038] S6: The leaf protein precipitate obtained in step S5 is placed in a -60°C freeze dryer and freeze-dried for 12 hours to obtain a leaf protein product.

[0039] Example 3

[0040] The present embodiment provides a method for extracting fresh Moringa leaf protein by adding molasses and fermenting, comprising the following steps:

[0041] S1: Wash the freshly picked Moringa leaves with clean water to remove impurities such as rotten leaves and sand; then cut the washed Moringa leaves into small pieces of 1-2 cm;

[0042] S2: placing the pretreated fresh Moringa leaves into a pulper, adding water to the pulper according to a solid-liquid ratio of 1:13.5, and pulping at a rate of 500 r / min for 3 min at 25° C. to obtain freshly squeezed leaf pulp;

[0043] S3: filtering the freshly squeezed leaf pulp in step S2 through a 300-mesh nylon mesh filter bag to separate the filter residue and the filtrate; cleaning the pulper, mixing the cleaning liquid with the filter residue and filtering again through a 300-mesh nylon mesh filter bag, repeating 3 times, and collecting all the filtrate;

[0044] S4: adding molasses accounting for 7% of the mass of the filtrate to the filtrate collected in step S3; stirring for 3 minutes at 25°C until uniform to obtain a mixed solution; and then placing in a constant temperature incubator at 30°C for anaerobic fermentation for 38 hours to obtain a fermentation liquid;

[0045] S5: placing the fermentation liquid in step S6 in a centrifuge, centrifuging at 5000 r / min for 15 min to separate the leaf protein precipitate and the supernatant, and discarding the supernatant to obtain the leaf protein precipitate;

[0046] S6: The leaf protein precipitate obtained in step S5 is placed in a -60°C freeze dryer and freeze-dried for 12 hours to obtain a leaf protein product.

[0047] Comparative Example 1

[0048] The difference between this comparative example and Example 1 is that in step S2, Moringa leaf dry powder is used instead of fresh Moringa leaves, and the other steps are the same.

[0049] Comparative Example 2

[0050] The difference between this comparative example and Example 1 is that no honey is added for modification in step S4, and the other steps are the same.

[0051] Comparative Example 3

[0052] The difference between this comparative example and Example 1 is that in step S5, the mixture is placed in a constant temperature incubator at 20° C. for anaerobic fermentation for 20 h, and the other steps are the same.

[0053] Comparative Example 4

[0054] The difference between this comparative example and Example 1 is that in step S7, freeze drying is performed in a -100°C freeze dryer for 5 hours, and the other steps are the same.

[0055] Test method:

[0056] p(%)=[(w1×p2) / (w2×p1×p3)]

[0057] Where: p: The percentage of extracted leaf protein in total protein (leaf protein extraction rate)

[0058] w1: dry weight of extracted leaf protein

[0059] w2: The quality of fresh Moringa leaves processed each time

[0060] p1: dry matter content in fresh Moringa samples

[0061] p2: Percentage of crude protein in the dry weight of leaf protein in the extract

[0062] p3: The percentage of crude protein in dry matter in fresh samples

[0063] The collected and extracted Moringa leaf protein concentrate was stored in a -20°C low-temperature refrigerator, and its crude protein content was determined by the Dumas combustion method.

[0064] Leaf protein extraction rate = (grams of protein in leaf protein / grams of protein in fresh sample) × 100%

[0065] Leaf protein yield = (leaf protein dry weight in grams obtained from fresh sample / fresh sample in grams) × 100%

[0066] The extraction rates of the various embodiments and comparative examples are shown in Table 1.

[0067] Table 1 Performance test results

[0068] test Leaf protein extraction rate % Leaf protein yield% Crude protein content% Example 1 58.37 6.9 43.5 Example 2 58.61 7.1 44.9 Example 3 59.55 7.6 46.1 Comparative Example 1 47.62 3.7 38.1 Comparative Example 2 48.21 3.9 38.5 Comparative Example 3 50.33 4.3 40.1 Comparative Example 4 49.75 4.1 39.3

[0069] Table 2 Response surface experimental design and results

[0070]

[0071]

[0072] Table 3 Analysis of variance of regression model

[0073]

[0074] Note: * indicates significant difference (P<0.05); ** indicates extremely significant difference (P<0.01).

[0075] From the above performance test results, it can be seen that the leaf protein extraction rate, leaf protein yield and crude protein content of Examples 1-3 are all high; in particular, the comprehensive performance of Example 3 is the most outstanding. However, since Comparative Examples 1-4 do not adopt the necessary technical solutions, their corresponding performance tests are significantly worse than those of the Examples. The above experimental results further prove the importance of the technical solutions defined in the present invention for its technical effects.

[0076] The above is a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for extracting fresh Moringa leaf protein by adding molasses and fermenting, characterized in that: The steps include: S1: Pre-treatment of fresh Moringa leaves; S2: placing the pre-treated fresh Moringa leaves into a pulping machine, adding water to pulp, and obtaining freshly squeezed leaf pulp; S3: filtering the freshly squeezed leaf pulp in step S2 through a nylon mesh filter bag to separate the filter residue and the filtrate; cleaning the pulper, mixing the cleaning liquid with the filter residue and filtering through a nylon mesh filter bag, repeating this process three times, and collecting all the filtrate; S4: adding molasses to the filtrate collected in step S3, mixing evenly to obtain a mixed solution; then placing the mixed solution in a constant temperature incubator at 25-45° C. for anaerobic fermentation for 35-45 hours to obtain a fermentation liquid; S5: placing the fermentation liquid in step S4 in a centrifuge, centrifuging to separate the leaf protein precipitate and the supernatant, and discarding the supernatant to obtain the leaf protein precipitate; S6: The leaf protein precipitate obtained in step S5 is placed in a -80°C freeze dryer and freeze-dried for 12 hours to obtain a leaf protein product.

2. A method for extracting fresh Moringa leaf protein by adding molasses fermentation according to claim 1, characterized in that, The pretreatment process of step S1 is: washing the picked fresh Moringa leaves with clean water to remove impurities such as rotten leaves, mud and sand; then cutting the washed fresh Moringa leaves into small segments of 1-2 cm.

3. A method for extracting fresh Moringa leaf protein by adding molasses fermentation according to claim 1, characterized in that, In step S2, the solid-liquid ratio of fresh Moringa leaves to water is 1:12-15; and the pulping is performed at a rate of 500 r / min for 2-3 min at 25° C.

4. A method for extracting fresh Moringa leaf protein by adding molasses fermentation according to claim 1, characterized in that, The pore size of the nylon mesh in step S3 is 300 meshes.

5. A method for extracting fresh Moringa leaf protein by adding molasses fermentation according to claim 1, characterized in that, The molasses in step S4 is sugarcane molasses; the molasses accounts for 5-10% of the mass of the filtrate; and the molasses is manually stirred for 3 minutes at 25°C until uniform.

6. A method for extracting fresh Moringa leaf protein by adding molasses fermentation according to claim 1, characterized in that, In step S5, centrifuge at 4500-6000 r / min for 10-20 min.

7. A method for extracting fresh Moringa leaf protein by adding molasses fermentation according to claim 1, characterized in that, In step S6, the leaf protein precipitate is placed in a -60°C freeze dryer and freeze-dried for 12 hours.

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