A bio-enzymatic preparation method for a macadamia nut plant protein milk beverage and its product.

Abstract

This invention discloses a bio-enzymatic preparation method for macadamia nut plant protein milk beverage and its product. The method includes mixing enzymatically hydrolyzed macadamia nut protein slurry, deionized water, macadamia nut oil diglycerides, skim milk, monoglycerides, sucrose esters, xanthan gum, carrageenan, sodium alginate, and white sugar; adjusting the pH to 6.5–7; homogenizing under high pressure; and sterilizing to obtain the macadamia nut plant protein milk beverage. This invention utilizes α-amylase and cellulase to synergistically hydrolyze insoluble macromolecules in macadamia nut meal, thereby increasing the solubility of the macadamia nut protein slurry and maximizing the release of small-molecule phenolic substances. Under optimal conditions, papain hydrolyzes the protein in the macadamia nut protein slurry to generate polypeptides with strong antioxidant and emulsifying properties, increasing the free radical scavenging ability and emulsifying stability of the enzymatically hydrolyzed macadamia nut protein slurry.

Description

Technical Field

[0001] This invention belongs to the field of plant-based beverage preparation, specifically relating to a bio-enzymatic preparation method for macadamia nut plant protein milk beverage and its product. Background Technology

[0002] Macadamia integrifolia is a tree nut native to Queensland and subtropical rainforests. Macadamia kernels are highly nutritious, typically containing 70-80% oil and 7-10% crude protein, making them a good source of both. Potassium content is the highest in most macadamia nuts, reaching 0.32%–0.43%, followed by magnesium at 0.16%–0.20%. Phosphorus, calcium, manganese, zinc, iron, and copper are also present in relatively high levels. In addition to these nutrients, macadamia nuts also contain phenolic acids and phytic acid.

[0003] Macadamia nut oil is primarily composed of monounsaturated fatty acids (MUFAs), with oleic acid being the main component and palmitoleic acid the secondary component. Linoleic acid, a polyunsaturated fatty acid, is the second largest component. Palmitoleic acid, an ω-7 monounsaturated fatty acid, possesses various biological functions. Studies have shown that palmitoleic acid can reduce inflammation in tissues with high metabolic rates; it can inhibit the expression of melanin-producing enzymes, exhibiting a certain skin-whitening effect; and it can also serve as a raw material for the development of lipid-based therapeutic drugs. Therefore, the high proportion of oleic and palmitoleic acids in macadamia nut oil has significant application value in lowering plasma total cholesterol and LDL cholesterol levels, preventing inflammation, and preventing cardiovascular diseases. Furthermore, macadamia nut oil also contains abundant tocopherols, tocotrienols, and squalene, which play an important role in promoting human health.

[0004] Currently, macadamia nuts are mainly sold and consumed as nuts, with few reports on their more value-added processed products. Due to their rich nutritional content and pleasant taste, processing them into plant-based protein milk beverages could significantly increase their added value. Existing reports on macadamia nut plant-based protein milk beverages all involve using macadamia kernels as raw materials and directly employing a colloid mill to reduce particle size. However, this method results in milk beverages with large particle sizes and poor stability. Furthermore, because macadamia nuts contain a high amount of oil, excessive consumption may increase the risk of obesity. Summary of the Invention

[0005] The purpose of this section is to outline some aspects of embodiments of the present invention and to briefly describe some preferred embodiments. Simplifications or omissions may be made in this section, as well as in the abstract and title of this application, to avoid obscuring the purpose of these documents; however, such simplifications or omissions should not be construed as limiting the scope of the invention.

[0006] In view of the problems existing in the above and / or prior art, the present invention is proposed.

[0007] Therefore, the purpose of this invention is to overcome the shortcomings of the prior art and provide a bio-enzymatic preparation method for macadamia nut plant protein milk beverage.

[0008] To solve the above-mentioned technical problems, the present invention provides the following technical solution: a bio-enzymatic preparation method for a macadamia nut plant protein milk beverage, comprising,

[0009] Macadamia nuts are roasted, crushed, and then pressed to obtain macadamia nut oil and meal.

[0010] Macadamia nut meal was formulated into a macadamia protein slurry, which was then hydrolyzed with α-amylase and cellulase, followed by a reaction with protease to obtain an enzymatically hydrolyzed macadamia protein slurry.

[0011] Glyceryl ester intermediates were prepared by lipase-catalyzed hydrolysis of macadamia nut oil. The addition of glycerol was carried out in three stages, and each stage was thoroughly mixed by high-pressure homogenization.

[0012] After glycerol hydrolysis, the glycerol hydrolysis product was catalyzed by a pH-treated, fixed-conformation lipase Lipase G50 and reacted with fatty acids to obtain macadamia nut oil diglyceride.

[0013] The macadamia nut plant protein milk beverage is obtained by mixing enzymatically hydrolyzed macadamia nut protein slurry, deionized water, macadamia nut oil diglyceride, skim milk, monoglyceride, sucrose ester, xanthan gum, carrageenan, sodium alginate and white sugar, adjusting the pH to 6.5-7, homogenizing under high pressure, and sterilizing.

[0014] As a preferred embodiment of the preparation method described in this invention, the macadamia nuts are baked at a temperature of 110–120°C for 10–20 minutes until the moisture content is less than 5%.

[0015] As a preferred embodiment of the preparation method described in this invention, the step of preparing macadamia nut meal into macadamia protein paste includes,

[0016] Macadamia nut meal was crushed, passed through a 100-mesh sieve, and mixed with deionized water at a material-to-liquid ratio. The mixture was stirred and sheared to obtain macadamia nut protein slurry, wherein the material-to-liquid ratio was 1:4 (m:v).

[0017] As a preferred embodiment of the preparation method described in this invention, the addition of α-amylase and cellulase hydrolysis includes,

[0018] Adjust the pH of the macadamia nut protein slurry to 6, add α-amylase at a concentration of 60-180 U / g of the substrate and cellulase at a concentration of 300-800 U / g of the substrate, and react in a water bath at 60°C for 2-4 hours.

[0019] As a preferred embodiment of the preparation method described in this invention, the protease addition reaction is wherein the protease is papain, the reaction pH is 6, the protease addition amount is 150 U / g of substrate by mass percentage, the reaction temperature is 50°C, and the reaction time is 2-3 h.

[0020] In a preferred embodiment of the preparation method described in this invention, the lipases in the glycerol hydrolysis reaction include Lipase AY-30SD, Lipase R, and Lipase DF-15;

[0021] In the glycerol hydrolysis reaction, the amount of monoglyceride added is 0.5-1.5%, the amount of lipase added is 5-10 wt%, the stirring speed is 600-800 rpm, and the reaction temperature is 50-70℃;

[0022] In the glycerol hydrolysis reaction, glycerol is added in three stages. In the first stage, 5-20% glycerol is added, homogenized at 20-40 MPa for 3-8 min, and reacted for 0.5-1.5 h. In the second stage, 20-40% glycerol is added, homogenized at 20-40 MPa for 3-8 min, and reacted for 0.5-1.5 h. In the third stage, 40-75% glycerol is added, homogenized at 20-40 MPa for 3-8 min, and reacted for 4-9 h.

[0023] As a preferred embodiment of the preparation method described in this invention, after the glycerolysis reaction is completed, the system temperature is heated to 80-90°C and stirred at a speed of 30-80 rpm for 0.5-1.5 h.

[0024] As a preferred embodiment of the preparation method described in this invention, the pH-treated immobilized glycerol lipase Lipase G50 comprises dissolving Lipase G50 in a buffer solution with a pH of 5.5 and then fixing its conformation by vacuum drying.

[0025] The conditions for the esterification reaction are as follows: the molar ratio of free fatty acids to the glycerol backbone of glycerol ester is 1-4:1, the temperature is 30-50℃, the stirring speed is 600-800 rpm, the enzyme dosage is 4-8 wt%, the vacuum degree is 0.1 MPa, and the reaction time is 8-16 h.

[0026] In a preferred embodiment of the preparation method described in this invention, the following components are present by weight percentage: 15-30% enzymatically hydrolyzed macadamia nut protein slurry, 25-45% deionized water, 6-12% macadamia nut oil diglyceride, 20-30% skim milk, 0.05-0.15% monoglyceride, 0.05-0.15% sucrose ester, 0.01-0.03% xanthan gum, 0.01-0.03% carrageenan, 0.01-0.03% sodium alginate, and 6-9% white sugar.

[0027] Another objective of this invention is to overcome the shortcomings of the prior art and provide a macadamia nut plant protein milk beverage prepared by a bio-enzymatic method.

[0028] Beneficial effects of this invention:

[0029] (1) This invention utilizes α-amylase and cellulase to synergistically hydrolyze insoluble macromolecules in macadamia nut meal, thereby increasing the solubility of macadamia nut protein slurry and maximizing the release of small molecule phenolic substances. Under optimal conditions, papain is used to hydrolyze the protein in macadamia nut protein slurry to generate polypeptides with strong antioxidant and emulsifying properties, thereby increasing the free radical scavenging ability and emulsification stability of enzymatically hydrolyzed macadamia nut protein slurry.

[0030] (2) In the preparation of macadamia nut oil diglyceride, the present invention utilizes a method of adding monoglycerides and adding glycerol stepwise coupled with high-pressure homogenization to increase the compatibility of glycerol with oil and improve the reaction efficiency. At the same time, the optimal catalytic conformation of Lipase G50 is fixed by pH treatment, which improves the esterification activity of Lipase G50. The lipase is then used as a catalyst to catalyze the esterification reaction between glycerol hydrolysis products and free fatty acids, so that monoglycerides are converted into diglycerides. This greatly increases the yield of diglycerides in the product and reduces the generation of harmful substances. Detailed Implementation

[0031] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to the examples in the specification.

[0032] Many specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be practiced in other ways different from those described herein, and those skilled in the art can make similar extensions without departing from the spirit of the invention. Therefore, the invention is not limited to the specific embodiments disclosed below.

[0033] Secondly, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in different places in this specification does not necessarily refer to the same embodiment, nor is it a single or selective embodiment that is mutually exclusive with other embodiments.

[0034] In this invention, α-amylase was purchased from Qingdao Haiweisen Biotechnology Co., Ltd., with an enzyme activity of 6000 U / g; cellulase was purchased from Qingdao Haiweisen Biotechnology Co., Ltd., with an enzyme activity of 100000 U / g; papain was purchased from Shanghai Xiangqi Biotechnology Co., Ltd., with an enzyme activity of 100000 U / g; alkaline protease was purchased from Jiangsu Maisheng Biotechnology Co., Ltd., with an enzyme activity of 50000 U / g; trypsin was purchased from Shanghai Xianghong Biotechnology Co., Ltd., with an enzyme activity of 4000 U / g; and neutral protease was purchased from Jiangsu Jiayuancheng Biotechnology Co., Ltd., with an enzyme activity of 20000 U / g.

[0035] (1) Determination of peptide content in hydrolysate: The gradient dilution method was used to prepare standard solutions of Gly-Gly-Tyr-Arg tetrapeptide at concentrations of 0.0, 0.2, 0.4, 0.8, 1.2, 1.4, 1.6 and 1.8 mg / mL using 5% trichloroacetic acid. 6.0 mL of each standard solution was added to 4.0 mL of biuret reagent, mixed in a vortex mixer, allowed to stand for 20 min, and then centrifuged at 4000 r / min for 10 min. The absorbance of the supernatant was measured at 540 nm (with the first tube as a blank control). A standard curve was established with peptide concentration as the x-axis (mg / mL) and absorbance as the y-axis.

[0036] Add 2.5 mL of walnut hydrolysate to 2.5 mL of 10% trichloroacetic acid, mix well in a vortex mixer, let stand for 20 min, centrifuge at 4000 r / min for 10 min, transfer the supernatant to a 50 mL volumetric flask and dilute to volume with 5% trichloroacetic acid. Take 6.0 mL of the above solution and place it in another test tube, add 4.0 mL of biuret reagent, mix well and let stand for 20 min, then centrifuge at 4000 r / min for 10 min. Take the supernatant and measure the absorbance at 540 nm. Calculate the peptide concentration in the hydrolysate by referring to the standard curve.

[0037] (2) Extraction of polyphenols: The enzymatically treated sample and the control sample were mixed thoroughly with a 1% (v / v) HCl-methanol solution at a ratio of 1:25 (m / V) and allowed to stand at 25°C for 24 h. The mixture was then centrifuged at 4000×g for 10 min, and the supernatant was collected. The precipitate was further mixed with a 1% HCl-methanol solution at a ratio of 1:25 and allowed to stand at 25°C for 24 h. The mixture was then centrifuged at 4000×g for 10 min, and the extracts were combined and evaporated to dryness at 40°C. The resulting extract was then dissolved in 3 mL of methanol and stored at -20°C for subsequent analysis.

[0038] (3) Determination of total polyphenol content: Pipette 0.1 mL of moderately diluted sample solution into a 5 mL test tube, then add 0.5 mL of 0.2 mol / L Lolin-phenol reagent and 0.8 mL of 7.5% Na2CO3 reagent in sequence, mix well, and let stand in the dark at 25℃ for 30 min before measuring the absorbance at a wavelength of 765 nm. The results are expressed as the total polyphenol content in each mL of almond milk equivalent to the amount of gallic acid (GAE), with the unit being μmol GAE / mL almond milk.

[0039] (4) Determination of DPPH free radical scavenging rate: Mix 500 μL of moderately diluted polyphenol extract with 0.3 mL of 0.6 mmol / L DPPH-ethanol solution, shake well, and react in a 25℃ water bath in the dark for 30 min. Zero the sample with methanol and measure the absorbance at 517 nm. Prepare a 0-2000 μmol / L Trolox solution to replace the sample for a standard curve. The results are expressed as the amount of Trolox equivalent to each gram of almond milk (μmol TE / mL almond milk).

[0040] (5) Centrifugal sedimentation rate: Accurately weigh a certain amount of sample and place it in a centrifuge tube. Centrifuge at 5000 r / min for 20 min, discard the supernatant, invert the centrifuge tube containing the sediment for 30 min, accurately weigh the sediment mass, and calculate the centrifugal sedimentation rate (SR). Each sample is measured in 3 parallel trials, and the average value is taken. The smaller the centrifugal sedimentation rate, the better the stability of the beverage.

[0041] SR / % = (M1 / M2) × 100

[0042] In the formula: M1 is the mass of the precipitate after centrifugation, g; M2 is the mass of the sample before centrifugation, g.

[0043] Determination of Lipase G50 esterification activity: Glycerol and oleic acid were added to a batch reactor at a molar ratio of 1:1, followed by the addition of 4% Lipase G50. The reaction was carried out at 30°C and a stirring rate of 600 rpm for 1 hour. The initial enzyme activity was assessed based on the esterification rate (%) of fatty acids in the system after 1 hour of reaction.

[0044] The sensory evaluation criteria in this invention are shown in the table below:

[0045]

[0046]

[0047] Example 1

[0048] (1) The macadamia nuts are baked at 100℃ for 20 minutes until the moisture content is less than 5%. After shelling, they are pressed by a hydraulic press to obtain macadamia nut oil and macadamia nut meal.

[0049] Macadamia nut pulp was pulverized using a pulverizer, passed through a 100-mesh sieve, and mixed with deionized water at a material-to-liquid ratio of 1:4 (m:v). The mixture was then stirred and sheared to obtain macadamia nut protein slurry.

[0050] The pH of the macadamia nut protein slurry was adjusted to 6, and α-amylase with a substrate concentration of 120 U / g and cellulase with a substrate concentration of 300 U / g were added. The mixture was then in a water bath at 60°C for 2 hours to degrade macromolecular starch and insoluble dietary fiber, thereby improving the overall emulsification stability of the protein slurry and further releasing small molecules such as polyphenols that are bound to macromolecules such as starch and dietary fiber, resulting in enzymatically hydrolyzed macadamia nut protein slurry 1.

[0051] Subsequently, the pH of the protein slurry was adjusted to 5, 6, 7, 8 and 9, respectively, and papain, alkaline protease, trypsin and neutral protease of 100 U / g were added to the slurry for restricted enzymatic hydrolysis. The reaction temperature was 60℃ and the reaction time was 2h to obtain enzymatically hydrolyzed macadamia nut protein slurry 2. The content of polyphenols and polypeptides, free radical scavenging ability and centrifugation precipitation rate in the protein slurry at different pH values ​​were analyzed to investigate the action characteristics of different enzymes.

[0052] Table 1. Indicators of macadamia nut protein slurry hydrolyzed by different proteases at different pH values.

[0053]

[0054] Because the proteins in macadamia nuts denature after being roasted at high temperatures, their solubility decreases. By using restricted hydrolysis with proteases to break down some of the proteins into peptides, the solubility is improved. Furthermore, different proteases produce peptides with varying free radical scavenging abilities and emulsifying activities.

[0055] By observing the hydrolysis of macadamia nut protein by different proteases at different pH values, it was found that at pH 6, the macadamia nut protein slurry hydrolyzed by papain had the highest polypeptide content, the highest free radical scavenging ability, and the lowest centrifugal precipitation rate. At the same time, due to its better hydrolysis effect, more polyphenols were released, which further improved the free radical scavenging rate of the protein slurry.

[0056] (2) Papain was added to the enzymatically hydrolyzed macadamia nut protein slurry 1 at pH 6, with 50 U / g, 100 U / g, 150 U / g, and 200 U / g of the material respectively. The reaction temperature was 60℃ and the reaction time was 2h to obtain enzymatically hydrolyzed macadamia nut protein slurry 2. The content of polyphenols and polypeptides, free radical scavenging ability and centrifugation precipitation rate in the protein slurry were analyzed under different enzyme addition amounts.

[0057] Table 2. Indicators of papain-hydrolyzed macadamia nut protein slurry under different enzyme dosages.

[0058]

[0059] By analyzing various indicators of papain-hydrolyzed macadamia nut protein slurry under different enzyme dosages, it was found that when the enzyme dosage was 150 U / g of material, the resulting enzymatically hydrolyzed macadamia nut protein slurry 2 had the highest polypeptide content, the highest free radical scavenging ability, and also a relatively high polyphenol content, while the centrifugal sedimentation rate was relatively low.

[0060] Enzymatically hydrolyzed macadamia nut protein slurry 1 was prepared by adding 150 U / g of papain to the material at pH 6 and reacting at temperatures of 40℃, 50℃, 60℃ and 70℃ for 2 h to obtain enzymatically hydrolyzed macadamia nut protein slurry 2. The content of polyphenols and peptides, free radical scavenging ability and centrifugation precipitation rate in the protein slurry at different temperatures were analyzed.

[0061] Table 3. Indicators of papain-hydrolyzed macadamia nut protein slurry at different temperatures

[0062]

[0063] By analyzing various indicators of papain hydrolyzed macadamia nut protein slurry at different temperatures, it was found that the highest peptide content, free radical scavenging ability, and polyphenol content, and the lowest centrifugal precipitation rate were obtained in the hydrolyzed macadamia nut protein slurry 2 at a temperature of 50℃. This indicates that the papain reaction at a temperature of 50℃ has the best reaction state.

[0064] (3) The enzymatically hydrolyzed macadamia nut protein slurry 1 was subjected to a pH of 6 and 150 U / g of papain was added to the material. The mixture was reacted at 50°C for 1 h, 2 h, 3 h and 4 h respectively to obtain the enzymatically hydrolyzed macadamia nut protein slurry 2. The content of polyphenols and polypeptides, free radical scavenging ability and centrifugation precipitation rate in the protein slurry were analyzed at different reaction times.

[0065] Table 4. Indicators of papain-hydrolyzed macadamia nut protein slurry at different reaction times

[0066]

[0067] By analyzing various indicators of papain-hydrolyzed macadamia nut protein slurry at different reaction times, it was found that when the reaction time was 2-3 hours, the resulting enzymatically hydrolyzed macadamia nut protein slurry 2 had higher peptide content, higher free radical scavenging ability, higher polyphenol content, and lower centrifugal precipitation rate. This indicates that a reaction time of 2-3 hours is more appropriate. A reaction time shorter than 2 hours is insufficient, while a reaction time longer than 3 hours is excessive, leading to a large amount of peptide degradation, which in turn affects the product's free radical scavenging ability.

[0068] Therefore, in order to achieve better free radical scavenging rate and better emulsion stability in the enzymatically hydrolyzed macadamia nut protein slurry, the final enzymatic hydrolysis conditions were selected as follows: papain as catalyst, enzyme dosage of 150 U / g material, pH of 6, temperature of 50℃, and reaction time of 2-3 h.

[0069] (4) Add macadamia nut oil to the batch reactor, fill it with nitrogen for protection, raise the temperature to 60°C, add monoglyceride at a mass fraction of 1% of macadamia nut oil, stir to fully dissolve the monoglyceride in the macadamia nut oil, and add glycerol in three stages according to a molar ratio of glycerol to macadamia nut oil of 3:1.

[0070] In the first stage, add 10% glycerol and homogenize at 20 MPa for 8 minutes to fully mix the glycerol with the macadamia nut oil. Then add lipase Lipase AY-30SD at a rate of 5 wt% of the weight of the macadamia nut oil and start stirring at 800 rpm. React for 1 hour and then filter out the lipase.

[0071] In the second stage, 30% glycerol was added, and the mixture was homogenized at 20 MPa for 8 minutes. The lipase was then added back in and reacted for 1 hour with the stirring speed at 800 rpm. The lipase was then filtered out.

[0072] In the third stage, 60% glycerol was added, homogenized at 20 MPa for 8 min, and lipase was added for 4 h of reaction at a stirring speed of 800 rpm.

[0073] After the reaction was completed, the system temperature was heated to 90℃ and stirred slowly at 30 rpm for 1 hour to accelerate the separation of glycerol and glycerides. The glycerol was then recovered and used as a raw material for a new glycerolysis reaction.

[0074] The product contained 53.9% monoglycerides, 41.5% diglycerides, and 4.6% triglycerides.

[0075] (5) Lipase G50 exhibits substrate specificity, showing activity only towards monoglycerides and diglycerides, but not towards triglycerides. Utilizing Lipase G50 to catalyze the esterification reaction of monoglycerides with fatty acids in glyceride products to generate diglycerides can significantly increase the yield of diglycerides in the reaction. However, Lipase G50 has an optimal reaction pH value, at which it exhibits maximum reactivity. This utilizes the fact that biomolecular enzymes are flexible in aqueous solutions, rigid in insoluble solutions, and possess structural memory properties.

[0076] Therefore, by screening for the optimal pH value of Lipase G50 and removing moisture through vacuum drying, Lipase G50 can maintain its optimal reaction conformation. Thus, buffer solutions with pH values ​​of 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, and 8 were prepared, and 20% (w / w) of Lipase G50 was added to each solution. The mixture was thoroughly mixed to reach ion exchange equilibrium, filtered to obtain the lipase, and then vacuum dried for 12 hours to remove moisture. Its esterification activity was then analyzed.

[0077] Table 5. Esterification activity of Lipase G50 at different pH values

[0078] pH value Esterification activity (%) 4 9.6 4.5 16.3 5 23.2 5.5 25.5 6 23.2 6.5 17.3 7 14.5 7.5 13.6 8 10.1 Unprocessed 15.1

[0079] As shown in the table above, among the Lipase G50 with fixed conformation after pH treatment, the esterification activity of Lipase G50 with pH between 4.5 and 6.5 is higher than that of untreated Lipase G50, while the esterification activity of Lipase G50 treated with pH 5.5 is the highest.

[0080] Lipases are soft in aqueous solutions and rigid in non-aqueous solutions. By adjusting the pH to bring lipases to their optimal catalytic state in the early stages, and removing water from the lipases to give them a rigid structure, the lipases can maintain their catalytic activity, thereby increasing their catalytic activity in organic environments and thus improving reaction efficiency.

[0081] A mixture of glycerides was added to a batch reactor, and free macadamia nut oil fatty acids were added based on the amount of glycerides' glycerol backbone to carry out esterification. The molar ratio of free fatty acids to glycerides' glycerol backbone was 3:1. The temperature was raised to 40°C and stirring was started at 800 rpm. Lipase G50, a glycerides lipase whose conformation was fixed by pH 5.5 buffer, was added at 6 wt% of the substrate mass. The reactor was then evacuated to a vacuum of 0.1 MPa and reacted for 12 h to obtain the glycerides product.

[0082] After the reaction was completed, the lipase was removed, and the fatty acids were removed by molecular distillation. The molecular distillation conditions were: distillation temperature of 170℃, pressure of 4Pa, and condenser temperature of 30℃.

[0083] The product contained 0.6% monoglyceride, 93.7% diglyceride, 5.7% triglyceride, with a diglyceride yield of 93.7%, 1.24 mg / kg glycidyl ester, and 0.21 mg / kg chloropropanol ester.

[0084] (6) Formulation of macadamia nut plant protein milk beverage

[0085] Take 20% enzymatically hydrolyzed macadamia nut protein slurry, add 45% deionized water, 8% macadamia nut oil functional lipids, 20% skim milk, and a compound emulsifier including 0.1% molecularly distilled monoglycerides and 0.1% sucrose esters; thickeners including 0.02% xanthan gum, 0.02% carrageenan, 0.03% sodium alginate, and 6.73% white sugar. Stir at high speed in an emulsifier to ensure thorough mixing. Heat the prepared emulsion to 65℃ and homogenize twice at 30 MPa. Adjust the pH of the homogenized emulsion and observe its stability at different pH values.

[0086] Table 6. Characteristics of macadamia nut plant protein milk beverage at different pH values

[0087]

[0088] Therefore, at a pH of 6.5-7, macadamia nut plant protein milk beverage has good sensory characteristics and a low centrifugal sedimentation rate, which means it has good stability.

[0089] The macadamia nut plant protein milk beverage is injected into an ultra-high temperature instantaneous sterilizer and sterilized at 135°C for 4 seconds before packaging.

[0090] Example 2

[0091] (1) The macadamia nuts are baked at 110°C for 15 minutes until the moisture content is less than 5%. After shelling, they are pressed by a hydraulic press to obtain macadamia nut oil and macadamia nut meal.

[0092] Macadamia nut pulp was pulverized using a pulverizer, passed through a 100-mesh sieve, and mixed with deionized water at a material-to-liquid ratio of 1:4 (m:v). The mixture was then stirred and sheared to obtain macadamia nut protein slurry.

[0093] The pH of the macadamia nut protein slurry was adjusted to 6, and α-amylase with a substrate concentration of 180 U / g and cellulase with a substrate concentration of 500 U / g were added. The mixture was then in a water bath at 60°C for 3 hours to degrade macromolecular starch and insoluble dietary fiber, thereby improving the overall emulsification stability of the protein slurry and further releasing small molecules such as polyphenols that are bound to macromolecules such as starch and dietary fiber, resulting in enzymatically hydrolyzed macadamia nut protein slurry 1.

[0094] Subsequently, the pH of the protein slurry was adjusted to 6, and 150 U / g of papain was added to each slurry. The reaction temperature was 50℃ and the reaction time was 2.5 h to obtain enzymatically hydrolyzed macadamia nut protein slurry 2. The content of polyphenols and peptides, free radical scavenging ability and centrifugation precipitation rate in the protein slurry were analyzed under different enzyme addition amounts.

[0095] Table 7. Indicators of Enzymatically Hydrolyzed Macadamia Nut Protein Pulp 2

[0096]

[0097] (2) Preparation of macadamia nut oil diglyceride

[0098] Macadamia nut oil was added to a batch reactor, nitrogen gas was introduced for protection, the temperature was raised to 70°C, monoglycerides were added at a mass fraction of 1.5% of macadamia nut oil, and the mixture was stirred to fully dissolve the monoglycerides in the macadamia nut oil. Glycerin was added in three stages at a molar ratio of glycerol to macadamia nut oil of 2:1.

[0099] In the first stage, add 5% glycerol and homogenize at 30 MPa for 5 minutes to fully mix the glycerol with the macadamia nut oil. Then add lipase DF-15 at a weight of 10 wt% of the macadamia nut oil and start stirring at 700 rpm. React for 0.5 hours and then filter out the lipase.

[0100] In the second stage, 20% glycerol was added, and the mixture was homogenized at 30 MPa for 5 minutes. The lipase was then added back in and reacted for 0.5 hours with a stirring speed of 700 rpm. The lipase was then filtered out.

[0101] In the third stage, 75% glycerol was added, homogenized at 30 MPa for 5 min, and lipase was added for 9 h of reaction at a stirring speed of 700 rpm.

[0102] After the reaction was completed, the system temperature was heated to 85℃ and stirred slowly at 50 rpm for 1.5 h to accelerate the separation of glycerol and glycerides, and the glycerol was recovered as a raw material for new glycerolysis reaction.

[0103] The product contained 50.4% monoglycerides, 43.9% diglycerides, and 5.7% triglycerides.

[0104] (3) Lipase G50 exhibits substrate specificity, showing activity only towards monoglycerides and diglycerides, but not towards triglycerides. Utilizing Lipase G50 to catalyze the esterification reaction of monoglycerides with fatty acids in glyceride products to generate diglycerides can significantly increase the yield of diglycerides in the reaction. However, Lipase G50 has an optimal reaction pH value, at which it exhibits maximum reactivity. Biomolecular enzymes are flexible in aqueous solutions but rigid in insoluble solutions, and possess structural memory properties. Therefore, by screening for the optimal pH value of Lipase G50 and removing moisture through vacuum drying, Lipase G50 can maintain its optimal reaction conformation.

[0105] Lipase G50 was dissolved in a pH 5.5 buffer solution and vacuum-dried to fix its conformation, yielding pH-treated and fixed-conformation Lipase G50. A mixture of glycerides was added to a batch reactor, and free macadamia nut oil fatty acids were added based on the glycerol backbone content of the glycerides for esterification. The molar ratio of free fatty acids to the glycerol backbone of the glycerides was 4:1. The temperature was raised to 50°C, and stirring was initiated at 700 rpm. The pH 5.5-treated and fixed-conformation glyceryl lipase Lipase G50 was added at 4 wt% of the substrate mass. Vacuum was applied at 0.1 MPa, and the reaction was carried out for 8 hours to obtain the glyceride product.

[0106] After the reaction, lipase was removed, and fatty acids were removed by molecular distillation. The molecular distillation conditions were: distillation temperature 170℃, pressure 4 Pa, and condenser temperature 30℃. The obtained product contained 0.8% monoglyceride, 92.9% diglyceride, and 6.3% triglyceride, with a diglyceride yield of 92.9%, 1.21 mg / kg glycidyl ester content, and 0.25 mg / kg chloropropanol ester content.

[0107] (4) Preparation of macadamia nut plant protein milk beverage

[0108] Take 30% enzymatically hydrolyzed macadamia nut protein slurry 2, add 25% deionized water, 6% macadamia nut oil functional lipids, 30% skim milk, and a compound emulsifier including 0.15% molecularly distilled monoglyceride and 0.05% sucrose ester, and thickeners including 0.01% xanthan gum, 0.03% carrageenan, 0.02% sodium alginate and 8.74% white sugar. Stir at high speed in an emulsifier to make it fully mixed. Heat the prepared emulsion to 65°C and homogenize it twice at 30 MPa.

[0109] Adjust the pH value to 7, inject the macadamia nut plant protein milk beverage into an ultra-high temperature instantaneous sterilizer, sterilize it at 135℃ for 4 seconds, and then package it.

[0110] Example 3

[0111] (1) Macadamia nuts were roasted at 120℃ for 10 minutes until the moisture content was less than 5%. After shelling, they were pressed by a hydraulic press to obtain macadamia nut oil and macadamia nut meal. The macadamia nut meal was crushed using a pulverizer, passed through a 100-mesh sieve, and mixed with deionized water at a material-to-liquid ratio of 1:4 (m:v). The mixture was stirred and sheared to obtain macadamia nut protein slurry.

[0112] The pH of the macadamia nut protein slurry was adjusted to 6, and α-amylase with a substrate concentration of 60 U / g and cellulase with a substrate concentration of 800 U / g were added. The mixture was then in a water bath at 60°C for 4 hours to degrade macromolecular starch and insoluble dietary fiber, thereby improving the overall emulsification stability of the protein slurry and further releasing small molecules such as polyphenols that are bound to macromolecules such as starch and dietary fiber, resulting in enzymatically hydrolyzed macadamia nut protein slurry 1.

[0113] Subsequently, the pH of the protein slurry was adjusted to 6, and 150 U / g of papain was added to each slurry. The reaction temperature was 50℃ and the reaction time was 3 h to obtain enzymatically hydrolyzed macadamia nut protein slurry 2. The content of polyphenols and peptides, free radical scavenging ability and centrifugation precipitation rate in the protein slurry were analyzed under different enzyme addition amounts.

[0114] Table 8. Indicators of Enzymatically Hydrolyzed Macadamia Nut Protein Pulp 2

[0115]

[0116] (2) Preparation of macadamia nut oil diglyceride

[0117] Macadamia nut oil was added to a batch reactor, nitrogen gas was introduced for protection, the temperature was raised to 50°C, monoglycerides were added at a mass fraction of 0.5% of macadamia nut oil, and the mixture was stirred to fully dissolve the monoglycerides in the macadamia nut oil. Glycerin was added in three stages at a molar ratio of 4:1 to macadamia nut oil.

[0118] In the first stage, add 20% glycerol and homogenize at 40 MPa for 3 minutes to fully mix the glycerol with the macadamia nut oil. Add lipase R at a rate of 8 wt% of the weight of the macadamia nut oil and start stirring at 600 rpm. React for 1.5 hours and then filter out the lipase.

[0119] In the second stage, 40% glycerol was added, and the mixture was homogenized at 40 MPa for 3 minutes. The lipase was then added back in and reacted for 1.5 hours with a stirring speed of 600 rpm. The lipase was then filtered out.

[0120] In the third stage, 40% glycerol was added, and the mixture was homogenized at 40 MPa for 3 minutes. Lipase was then added and reacted for 5 hours with a stirring rate of 600 rpm. After the reaction was completed, the system temperature was heated to 80°C, and the mixture was slowly stirred at 80 rpm for 0.5 hours to accelerate the separation of glycerol and glycerides. The glycerol was then recovered and used as a raw material for a new glycerolysis reaction.

[0121] The product contained 55.8% monoglycerides, 40.7% diglycerides, and 3.5% triglycerides.

[0122] Lipase G50, a metaglycerol lipase, exhibits substrate specificity, showing activity only towards monoglycerides and diglycerides, but not towards triglycerides. Utilizing Lipase G50 to catalyze the esterification reaction of monoglycerides with fatty acids in glycerol products to produce diglycerides can significantly increase the yield of diglycerides in the reaction. However, Lipase G50 has an optimal reaction pH, at which it exhibits maximum reactivity. Biomolecular enzymes are flexible in aqueous solutions but rigid in insoluble solutions, and possess structural memory properties. Therefore, by selecting the optimal pH for Lipase G50 and removing moisture through vacuum drying, its optimal reaction conformation can be maintained.

[0123] Lipase G50 was dissolved in a buffer solution with a pH of 5.5, and then vacuum dried to fix the conformation, resulting in pH-treated and fixed Lipase G50.

[0124] A mixture of glycerides was added to a batch reactor, and free macadamia nut oil fatty acids were added based on the amount of glycerides' glycerol backbone to carry out the esterification reaction. The molar ratio of free fatty acids to glycerides' glycerol backbone was 1:1. The temperature was raised to 30°C and stirring was started at a speed of 600 rpm. Lipase G50, a glycerides lipase whose conformation was fixed by pH 5.5 buffer, was added at a rate of 8 wt% of the substrate mass. The reactor was then evacuated to a vacuum of 0.1 MPa and reacted for 16 h to obtain the glycerides product.

[0125] After the reaction, the lipase was removed, and the fatty acids were removed by molecular distillation. The molecular distillation conditions were: distillation temperature of 170℃, pressure of 4Pa, and condenser temperature of 30℃.

[0126] The product contained 0.7% monoglyceride, 94.7% diglyceride, 4.6% triglyceride, with a diglyceride yield of 94.7%, 1.28 mg / kg glycidyl ester, and 0.29 mg / kg chloropropanol ester.

[0127] (3) Formulation of macadamia nut plant protein milk beverage

[0128] Take 15% enzymatically hydrolyzed macadamia nut protein slurry, add 40% deionized water, 12% macadamia nut oil functional lipids, 25% skim milk, and a compound emulsifier including 0.05% molecularly distilled monoglycerides and 0.15% sucrose esters; thickeners including 0.03% xanthan gum, 0.01% carrageenan, 0.01% sodium alginate, and 7.75% white sugar. Stir at high speed in an emulsifier to ensure thorough mixing. Heat the prepared emulsion to 65℃ and homogenize twice at 30MPa. Adjust the pH to 6.5, then inject the macadamia nut plant protein milk beverage into an ultra-high temperature instantaneous sterilizer and sterilize at 135℃ for 4 seconds before packaging.

[0129] The scores obtained from the sensory evaluation of the macadamia nut plant protein milk beverages prepared in Examples 1-3 are shown in the table below.

[0130] Table 9 Sensory evaluation scores and centrifugal sedimentation rates of macadamia nut plant protein milk beverages

[0131]

[0132] Comparative Example 1

[0133] Referring to Example 2, the baking temperature and time were changed, while other conditions remained the same as in Example 2. The specific differences are as follows:

[0134] (1) Preparation of enzymatically hydrolyzed macadamia nut protein slurry

[0135] Macadamia nuts are roasted at 90°C for 25 minutes until the moisture content is 8%. After shelling, they are pressed using a hydraulic press to obtain macadamia nut oil and macadamia nut meal.

[0136] Macadamia nut pulp was pulverized using a pulverizer, passed through a 100-mesh sieve, and mixed with deionized water at a material-to-liquid ratio of 1:4 (m:v). The mixture was then stirred and sheared to obtain macadamia nut protein slurry.

[0137] The pH of the macadamia nut protein slurry was adjusted to 6, and α-amylase with a substrate concentration of 180 U / g and cellulase with a substrate concentration of 500 U / g were added. The mixture was then in a water bath at 60°C for 3 hours to degrade macromolecular starch and insoluble dietary fiber, thereby improving the overall emulsification stability of the protein slurry and further releasing small molecules such as polyphenols that are bound to macromolecules such as starch and dietary fiber, resulting in enzymatically hydrolyzed macadamia nut protein slurry 1.

[0138] Subsequently, the pH of the protein slurry was adjusted to 6, and 150 U / g of papain was added to each slurry. The reaction temperature was 50℃ and the reaction time was 2.5 h to obtain enzymatically hydrolyzed macadamia nut protein slurry 2.

[0139] Macadamia nut oil diglycerides were prepared according to the process in Example 2, and a macadamia nut plant protein milk beverage was obtained by blending the macadamia nut plant protein milk beverage.

[0140] Sensory evaluation of the product is shown in the table below:

[0141]

[0142] Sensory evaluation revealed that, under these conditions, the flavor of the macadamia nut plant protein milk beverage was not strong in terms of nutty flavor, but rather in terms of milk flavor, which affected the overall flavor quality of the product.

[0143] Comparative Example 2

[0144] Referring to Example 3, in the preparation of macadamia nut oil diglycerides, instead of using pH-controlled conformation Lipase G50 for esterification, molecular distillation was used to directly remove fatty acids. The specific difference is as follows:

[0145] (2) Preparation of macadamia nut oil diglyceride

[0146] Macadamia nut oil was added to a batch reactor, nitrogen gas was introduced for protection, the temperature was raised to 50°C, monoglycerides were added at a mass fraction of 0.5% of macadamia nut oil, and the mixture was stirred to fully dissolve the monoglycerides in the macadamia nut oil. Glycerin was added in three stages at a molar ratio of 4:1 to macadamia nut oil.

[0147] In the first stage, add 20% glycerol and homogenize at 40 MPa for 3 minutes to fully mix the glycerol with the macadamia nut oil. Add lipase R at a rate of 8 wt% of the weight of the macadamia nut oil and start stirring at 600 rpm. React for 1.5 hours and then filter out the lipase.

[0148] In the second stage, 40% glycerol was added, and the mixture was homogenized at 40 MPa for 3 minutes. The lipase was then added back in and reacted for 1.5 hours with a stirring speed of 600 rpm. The lipase was then filtered out.

[0149] In the third stage, 40% glycerol was added, and the mixture was homogenized at 40 MPa for 3 minutes. Lipase was then added and reacted for 5 hours with a stirring rate of 600 rpm. After the reaction was completed, the system temperature was heated to 80°C, and the mixture was slowly stirred at 80 rpm for 0.5 hours to accelerate the separation of glycerol and glycerides. The glycerol was then recovered and used as a raw material for a new glycerolysis reaction.

[0150] The product contained 55.8% monoglycerides, 40.7% diglycerides, and 3.5% triglycerides.

[0151] Fatty acids were removed by single-stage molecular distillation under the following conditions: distillation temperature 170℃, pressure 4 Pa, and condenser temperature 30℃. The resulting product contained 54.5% monoglycerides, 41.9% diglycerides, and 3.6% triglycerides, with a diglyceride yield of 41.9%, glycidyl ester content of 1.96 mg / kg, and chloropropanol ester content of 0.52 mg / kg. Monoglycerides were then removed by two-stage molecular distillation under the following conditions: distillation temperature 200℃, pressure 2 Pa, and condenser temperature 25℃. The resulting product contained 89.7% diglycerides, 0.9% monoglycerides, 9.3% triglycerides, 5.16 mg / kg glycidyl ester content, and 1.14 mg / kg chloropropanol ester content.

[0152] Comparative Example 3

[0153] Referring to Example 2, the preparation of macadamia nut oil diglyceride did not employ the method of segmented addition and homogenization mixing of glycerol, but instead involved direct addition of glycerol for reaction. The difference lies in:

[0154] (2) Preparation of macadamia nut oil diglyceride

[0155] Macadamia nut oil was added to a batch reactor, nitrogen gas was introduced for protection, the temperature was raised to 70°C, monoglycerides were added at a mass fraction of 1.5% of macadamia nut oil, and the mixture was stirred to fully dissolve the monoglycerides in the macadamia nut oil. Glycerin was added at a molar ratio of glycerin to macadamia nut oil of 2:1.

[0156] Add lipase DF-15 at a rate of 10 wt% of the weight of macadamia nut oil, and start stirring at 700 rpm for 10 hours.

[0157] After the reaction was completed, the system temperature was heated to 85℃ and stirred slowly at 50 rpm for 1.5 h to accelerate the separation of glycerol and glycerides, and the glycerol was recovered as a raw material for new glycerolysis reaction.

[0158] The product contained 39.1% monoglycerides, 41.3% diglycerides, and 19.5% triglycerides. The high triglyceride content indicates that the reaction efficiency was significantly reduced in the system without homogeneous mixing.

[0159] Comparative Example 4

[0160] Under the conditions of Example 1, macadamia nut oil diglyceride was not prepared, but directly replaced with macadamia nut oil. The formulation of the macadamia nut plant protein milk beverage was the same as in Example 1. The stability of the macadamia nut plant protein milk beverage was determined, and the stability results were obtained by referring to the method for determining the centrifugal sedimentation rate.

[0161] The measurement results are shown in the table below.

[0162] Centrifugal sedimentation rate (%) Example 1 0.6 Comparative Example 4 1.2

[0163] It can be seen that when macadamia nut oil diglyceride is not prepared and is directly replaced with macadamia nut oil, the stability of the resulting dairy product decreases. Because diglyceride contains two fatty acid acyl groups and one hydroxyl group on its glycerol backbone, it possesses certain emulsifying properties. It works synergistically with other substances in the system to enhance the stability of the dairy beverage system.

[0164] Comparative Example 5

[0165] Under the conditions of Example 2, only enzymatically hydrolyzed macadamia nut protein slurry 1 was prepared, without preparing enzymatically hydrolyzed macadamia nut protein slurry 2. Other processes were the same as in Example 2. A milk beverage was prepared, and the stability of the macadamia nut plant protein milk beverage was determined. The stability results were obtained by referring to the method for determining the centrifugal sedimentation rate.

[0166] The measurement results are shown in the table below.

[0167] Centrifugal sedimentation rate (%) Example 2 0.5 Comparative Example 5 2.2

[0168] It can be seen that when the enzymatically hydrolyzed macadamia nut protein slurry 2 is not prepared, and the peptides are not added to generate the final beverage, the stability of the beverage is not good. Since the generated peptides have certain emulsifying properties and better solubility, they work together with other substances to improve the stability of the almond milk beverage emulsion system. At the same time, due to α-amylase, the cellulose hydrolysis process will release a certain amount of polyphenols. These polyphenols bind to proteins and reduce the solubility of proteins to a certain extent.

[0169] Comparative Example 6

[0170] Under the conditions of Example 2, macadamia nut meal was simply prepared into macadamia protein slurry without the addition of α-amylase or cellulase for hydrolysis. Instead, protease was directly added to generate polypeptides. Other processes were the same as in Example 2. A milk beverage was prepared, and the stability of the macadamia nut plant protein milk beverage was determined. The stability results were obtained by referring to the method for determining the centrifugation sedimentation rate.

[0171] The measurement results are shown in the table below.

[0172] Centrifugal sedimentation rate (%) Example 2 0.5 Comparative Example 6 2.4

[0173] It can be seen that when protease is added directly to generate peptides without the addition of α-amylase and cellulase for hydrolysis, the resulting beverage exhibits poor stability. Since some starch and insoluble dietary fiber present in macadamia nut meal negatively impact beverage stability, failure to hydrolyze them into soluble substances will reduce the beverage's stability.

[0174] Comparative Example 7

[0175] Under the conditions of Example 2, macadamia nut meal was formulated into macadamia protein slurry without the addition of α-amylase or cellulase for hydrolysis, and without the addition of protease to generate peptides. Other processes were the same as in Example 2. A milk beverage was prepared, and the stability of the macadamia nut plant protein milk beverage was determined. The stability results were obtained by referring to the method for determining the centrifugal sedimentation rate.

[0176] The measurement results are shown in the table below.

[0177] Centrifugal sedimentation rate (%) Example 2 0.5 Comparative Example 7 3.1

[0178] It can be seen that when macadamia nut meal is formulated into a macadamia protein slurry, without the addition of α-amylase and cellulase for hydrolysis, and without the addition of protease to generate peptides, the resulting beverage exhibits poor stability. This is because macadamia nut meal contains insoluble starch, dietary fiber, and protein, which have a significant negative impact on beverage stability, ultimately leading to reduced stability.

[0179] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all such modifications or substitutions should be covered within the scope of the present invention.

Claims

1. A bio-enzymatic preparation method for a macadamia nut plant protein milk beverage, characterized in that: include, Macadamia nuts are roasted, crushed, and then pressed to obtain macadamia nut oil and meal. Macadamia nut meal was formulated into macadamia nut protein slurry, hydrolyzed with α-amylase and cellulase, and then reacted with protease to obtain enzymatically hydrolyzed macadamia nut protein slurry. The addition of α-amylase and cellulase hydrolysis was carried out by adjusting the pH of the macadamia nut protein slurry to 6, adding 60-180 U / g of α-amylase and 300-800 U / g of cellulase, and reacting in a water bath at 60°C for 2-4 hours. The addition of protease was carried out by using papain, with a reaction pH of 6, an addition amount of 150 U / g of protease, a reaction temperature of 50°C, and a reaction time of 2-3 hours. Glyceryl ester intermediates were prepared by lipase-catalyzed glycerol hydrolysis of macadamia nut oil. The addition of glycerol was carried out in three stages, with each stage involving thorough mixing via high-pressure homogenization. The lipases used in the glycerol hydrolysis reaction included Lipase AY-30SD, Lipase R, and Lipase DF-15. The amount of monoglyceride added was 0.5–1.5%, the amount of lipase added was 5–10 wt%, the stirring speed was 600–800 rpm, and the reaction temperature was 50–70 °C. Glycerol was added in three stages: in the first stage, 5–20% glycerol was added, homogenized at 20–40 MPa for 3–8 min, and reacted for 0.5–1.5 h; in the second stage, 20–40% glycerol was added, homogenized at 20–40 MPa for 3–8 min, and reacted for 0.5–1.5 h; in the third stage, 40–75% glycerol was added, homogenized at 20–40 MPa for 3–8 min, and reacted for 4–9 h. After glycerol hydrolysis, the glycerol hydrolysis product is catalyzed by a pH-treated, fixed-conformation lipase, Lipase G50, in a fatty acid esterification reaction to obtain macadamia nut oil diglyceride with low levels of harmful substances. The pH-treated, fixed-conformation lipase, Lipase G50, is obtained by dissolving Lipase G50 in a pH 5.5 buffer solution, followed by vacuum drying to fix the conformation. The esterification reaction conditions are: a molar ratio of free fatty acids to the glycerol backbone of the glycerol ester is 1–4:1; the temperature is 30–50°C; the stirring speed is 600–800 rpm; the enzyme dosage is 4–8 wt%; the vacuum degree is 0.1 MPa; and the reaction time is 8–16 h. The macadamia nut oil diglyceride has a glycidyl ester content of less than 1.3 mg / kg and a chloropropanol ester content of less than 0.3 mg / kg. A macadamia nut plant protein milk beverage is obtained by mixing enzymatically hydrolyzed macadamia nut protein slurry, deionized water, macadamia nut oil diglycerides, skim milk, monoglycerides, sucrose esters, xanthan gum, carrageenan, sodium alginate, and white sugar, adjusting the pH to 6.5-7, homogenizing under high pressure, and sterilizing. The beverage comprises, by weight percentage of the raw materials: 15-30% enzymatically hydrolyzed macadamia nut protein slurry, 25-45% deionized water, 6-12% macadamia nut oil diglycerides, 20-30% skim milk, 0.05-0.15% monoglycerides, 0.05-0.15% sucrose esters, 0.01-0.03% xanthan gum, 0.01-0.03% carrageenan, 0.01-0.03% sodium alginate, and 6-8% white sugar.

2. The preparation method according to claim 1, characterized in that: The process involves baking macadamia nuts at a temperature of 110-120°C for 10-20 minutes until the moisture content is less than 5%.

3. The preparation method according to claim 1, characterized in that: The process of preparing macadamia nut meal into macadamia nut protein paste includes, Macadamia nut meal was crushed, passed through a 100-mesh sieve, and mixed with deionized water at a material-to-liquid ratio. The mixture was stirred and sheared to obtain macadamia nut protein slurry, wherein the material-to-liquid ratio was 1:4 (m:v).

4. The preparation method according to claim 1, characterized in that: After the glycerol hydrolysis reaction is completed, the system temperature is heated to 80~90℃ and stirred at 30~80 rpm for 0.5~1.5h.

5. The macadamia nut plant protein milk beverage prepared by any one of claims 1 to 4.

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