CN113951366A

CN113951366A - Processing method for preparing biological enzymolysis protein by using squid leftovers

Info

Publication number: CN113951366A
Application number: CN202111251923.0A
Authority: CN
Inventors: 王岳庆; 夏宇; 王昌友
Original assignee: Zhoushan Fushi Food Technology Co ltd
Current assignee: Zhoushan Fushi Food Technology Co ltd
Priority date: 2021-10-26
Filing date: 2021-10-26
Publication date: 2022-01-21

Abstract

A processing method for preparing biological enzymolysis protein by using squid leftovers comprises the steps of taking the squid leftovers as raw materials, performing protein denaturation by adopting a heat treatment mode, adding exogenous protease, determining papain and flavourzyme as the best protease in a subsequent enzymolysis process, obtaining the best enzymolysis conditions of the papain and flavourzyme by experiments, performing single-factor experiments by selecting three factors of air inlet temperature, feed material concentration and drying aid adding amount, determining the best experimental process of the three factors according to the wall sticking condition in the spraying process and the quality of hydrolyzed protein powder, performing orthogonal experiments on the basis to determine the best process conditions of spray drying, obtaining the hydrolyzed squid protein powder with better quality, having few wall sticking phenomena in the spray drying process, being difficult to generate caramelization reaction and Maillard reaction, and having moderate water content of the protein powder, the powder has better dispersibility and fluidity; has good color, no browning, fragrant squid taste and good quality.

Description

Processing method for preparing biological enzymolysis protein by using squid leftovers

Technical Field

The invention relates to the technical field of food processing, in particular to a processing method for preparing biological enzymolysis protein by using squid leftovers.

Background

The squid is an important ocean economic cephalopod, is widely distributed in various sea areas of the Atlantic ocean, the Indian ocean and the Pacific ocean, and is a domestic main ocean fishing variety and an aquatic product processing product. According to the ocean conference data of the Chinese fishery association, the squid has the advantages of high protein, low fat and low calorie, and the nutritive value of the squid is not inferior to that of beef and tuna. The Loligo chinensis Gray has the advantages of high protein, low fat, and low calorie. The fish is rich in various amino acids necessary for human body, and the composition of the necessary amino acids is close to that of the whole protein, so that the fish is a nutritional health-care aquatic product resource with good flavor. In addition, Loligo chinensis Gray also contains abundant flavor substances such as taurine, lysine, octocarnitine, betaine, noisy, nucleotides, saccharides, and trimethylamine oxide.

In the process of processing the squid into squid shreds, squid rings, squid flowers or seasoning squid products, about 15 to 20 percent of waste such as internal organs, epidermis and the like are generated, although the waste also has high nutritive value, only a few factories extract squid oil or prepare fish meal from the waste to be used as aquatic feeds. Most factories adopt a method of burying and discarding, which not only causes great waste of resources, but also causes environmental pollution. In fact, the wastes have higher utilization value, are rich in physiologically active substances such as protein, collagen and hydrolyzed polypeptide thereof and have the effects of resisting oxidation, tumors and the like.

Macromolecular proteins of aquatic animals such as squid leftovers and the like are hydrolyzed into short peptides and amino acids by an enzymolysis method, so that the squid polypeptide hydrolysate has multiple physiological functions of resisting oxidation, reducing blood pressure and the like, can be used as a nitrogen source of microorganisms, and can also be used for preparing hydrolyzed protein products so as to further improve the deep utilization of the squid. At present, squid processing is only limited to ice fresh, processing of dry products is low in product added value, domestic deep processing capacity is insufficient, the variety of products can be enriched through research of squid powder products, meanwhile, flavor peptides and functional peptides obtained through enzymolysis are beneficial to improvement of free amino acid and polypeptide content, the nutritional value of the products is improved, and absorption of a human body is facilitated.

Disclosure of Invention

In order to solve the technical defects in the prior art, the invention provides an intestinal tract flexible anastomosis stent.

The technical solution adopted by the invention is as follows: the processing method for preparing the biological enzymolysis protein by using the squid leftovers comprises the following steps:

(1) selection of raw materials: selecting fresh Loligo chinensis Gray or frozen Loligo chinensis Gray, and removing viscera;

(2) pulping: pulping the treated squid, and repeatedly pulping until the fineness is below 5 mm;

(3) rubber grinding: adding the squid enzymolysis liquid into a colloid mill, and controlling the gap to be 20-50 um;

(4) carrying out enzymolysis pretreatment: heat treating the pulped Loligo chinensis Gray at 50-90 deg.C for 15 min;

(5) and (3) carrying out enzymolysis on the squid protein: adding papain or flavourzyme into the pretreated raw materials to be used as exogenous protease for hydrolyzing squid protein for hydrolysis, controlling the temperature to be 45-50 ℃, adjusting the pH value of reaction liquid to be 6.0-7.0, controlling the hydrolysis time to be 2-4h, wherein the enzyme adding amount of the papain is 600U/g of squid, the enzyme adding amount of the flavourzyme is 500U/g of squid, and the substrate concentration is 8%;

(6) blending: adding fishy smell removing substance, and adjusting its content;

(7) homogenizing: homogenizing the mixed liquid;

(8) concentration: low-temperature vacuum concentration is adopted, and the concentration conditions are as follows: at 45-50 deg.C, vacuum degree of 10-13KPa, and concentrated vegetable liquid solid content of 55-60%;

(9) spray drying: drying by adopting a pressure type spray dryer, adding a spray drying auxiliary agent, controlling the pressure to be 12.5-15.0MPa, and controlling the air inlet temperature: 170 ℃ and 190 ℃, and the air exhaust temperature: the water content of the dried material is less than 5 percent at 70-80 ℃.

When the exogenous protease for hydrolyzing the squid protein is papain, the hydrolysis temperature is 50 ℃, and the pH value of the reaction solution is adjusted to be 7.0.

When the exogenous protease for hydrolyzing the squid protein is the flavor protease, the hydrolysis temperature is 45 ℃, and the pH value of the reaction solution is adjusted to 6.0.

The air inlet temperature during spray drying is 190 ℃.

The feed concentration of the feed liquid during spray drying is 15%.

During spray drying, the ratio of the squid solid matter serving as the spray drying assistant agent to the maltodextrin is 4: 1.

The treatment temperature of the enzymolysis pretreatment is 90 ℃.

The invention has the beneficial effects that: the invention provides a processing method for preparing biological enzymolysis protein by using squid leftovers, which takes the squid leftovers as raw materials, denaturizes the protein by adopting a heat treatment mode, adds exogenous protease, determines that papain and flavourzyme are used as the optimal protease in a subsequent enzymolysis process, obtains the optimal enzymolysis conditions of the papain and flavourzyme through experiments, has higher hydrolysis degree and protein content, performs single-factor experiments by selecting three factors of air inlet temperature, feed concentration and additive amount of a drying aid, and determines the optimal experimental process of the three factors according to the wall sticking condition in the spraying process and the quality of hydrolyzed protein powder. And performing orthogonal experiment on the basis to determine the optimal process condition of spray drying to obtain hydrolyzed squid protein powder with better quality, wherein the wall sticking phenomenon in the spray drying process is less, and caramelization reaction and Maillard reaction are not easy to occur. The water content of the protein powder is moderate, and the powder has better dispersibility and fluidity; has good color, no browning, fragrant squid taste and good quality.

Drawings

FIG. 1 is a graph showing the effect of pretreatment temperature on the degree of enzymatic hydrolysis.

FIG. 2 is a graph showing the effect of pretreatment time on the degree of enzymatic hydrolysis.

FIG. 3 is a graph showing the hydrolysis of fish protein by papain and flavourzyme in eggs.

FIG. 4 is a graph showing the effect of pH on the degree of squid proteolysis.

FIG. 5 is a graph of the effect of substrate concentration on the degree of proteolysis in fish.

FIG. 6 is a graph showing the effect of enzyme addition on enzymatic digestion of fish protein.

FIG. 7 is a graph showing the effect of inlet air temperature on water content and amino acid nitrogen content.

FIG. 8 is a graph showing the effect of feed concentration on water content and amino acid nitrogen content.

FIG. 9 is a graph showing the effect of maltodextrin on water content and amino acid nitrogen content.

Detailed Description

The technical solutions in the embodiments of the present invention will be described clearly and completely with reference to the accompanying drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be obtained by a person skilled in the art without any inventive step based on the embodiments of the present invention, are within the scope of the present invention.

Experimental Material

Squid leftovers; preparing an enzymolysis raw material: heating the minced Loligo chinensis Gray at 90 deg.C for 15min, mincing with tissue triturator, packaging, and storing at-20 deg.C. The experimental reagent is flavor enzyme; papain; pepsin; a neutral protease; (ii) trypsin; salicylic acid; dithiothreitol; sodium benzoate; boric acid. The experimental apparatus was a muffle furnace (two-remote fire technologies, ltd, beijing); ultraviolet spectrophotometer (Shimadzu corporation, Japan); low speed centrifuges (Beijing medical centrifuge Mill); electronic balances (Shanghaiengaceae balances); acidimeters (Shanghai Lei magnetic Instrument works).

Process flow

Frozen squid leftovers are ground into raw materials → unfreezing → water is added for even mixing and pulping and colloid milling → pretreatment → enzyme addition → hydrolysis is carried out by controlling proper factors such as time, temperature and the like → water bath at 95 ℃ for enzyme deactivation for 15min → cooling → centrifugation (9000r/min, 15min) → concentration → spray drying → protein powder.

Determination of the degree of hydrolysis

Preparation of OPA reagent: 9.550g of disodium tetraborate decahydrate and 250mg of SDS were completely dissolved in deionized water, 200mg of OPA was dissolved in 5ml of ethanol, the OPA solution was transferred to the above solution, 220mg of dithiothreitol was added to the solution, and finally, 250ml of deionized water was added to the solution. 5ml of the solution was aspirated and centrifuged at 920g for 10min, and 2ml of the supernatant was diluted to 50ml with deionized water as a sample solution.

Serine standard solution: 50mg of serine was made up to 500ml with deionized water. Four replicates were made for each sample, standard and blank. Adding 3ml of OPA reagent and 400uL of sample solution or serine standard solution into a 15ml test tube, starting timing after 5s of reaction, accurately reacting for 2min, and measuring the absorbance at 340 nm. The mean values of the standard and blank were used for calculation.

The degree of hydrolysis is calculated as follows: h/h_totle×100％

Wherein: h is the number of peptide bonds hydrolyzed; h is_totleThe total number of peptide bonds of an equal amount of protein.

Determination of squid enzymolysis process

Selection of the type of hydrolytic protease: five proteases, namely papain, neutral protease, flavourzyme, pepsin and trypsin, are selected to respectively hydrolyze the edible part of the squid. Controlling the enzymolysis condition as 5% (w/w) of substrate concentration, and comparing the hydrolysis effects of squid protein with the hydrolysis degree, bitter taste and clarification condition of hydrolysate as indexes, wherein the hydrolysis conditions are shown in Table 1.

TABLE 1 hydrolysis conditions for the various proteases

According to table 2, the hydrolysis of squid is known to be separately carried out by five proteases. Wherein when the squid protein is hydrolyzed by adopting neutral protease, pepsin and trypsin, the bitterness is heavier, and the hydrolysate is muddy in the pond. When the squid protein is subjected to enzymolysis by neutral protease and trypsin, the solubility of the protein is higher at higher concentration, and the hydrolysis degree is improved. And many macromolecular proteins and polypeptides form an emulsification system with a small amount of fat in the hydrolysate, and the hydrolysate is relatively muddy. Trypsin is expensive and less effective than papain and flavor enzymes, which are not selected for subsequent industrialization. When the squid protein is hydrolyzed by adopting the papain and the flavor enzyme respectively, the hydrolysis degree of the protein is higher, and meanwhile, the bitterness of the hydrolysate is weaker. According to the experimental results in the table 2 and the respective action characteristics of the five enzymes, the papain and the flavor enzyme are determined to be used for hydrolyzing the squid protein

TABLE 2 Effect of protease species on fish proteolysis

Determination of squid pretreatment conditions: the squid carcass consists of four layers of epidermis, two layers of endothelium and a muscle layer sandwiched between the epidermis and the endothelium. The epidermis is composed of tough and tenacious collagen fibers, the first and second layers have no reticular tissues with definite direction, the core is provided, and melanocytes are distributed in the second layer and between the two layers. The third layer is a reticular tissue similar to muscle, multinucleated. The fourth and inner layers are typically myofibrillar connective tissue. When exogenous protease is added to hydrolyze squid protein, the particularity and stability of the muscle layer structure and the enzyme cutting sites are occluded in protein molecules, so that the squid protein is difficult to hydrolyze by the protease, and pretreatment is required to denature the protein molecules. Protein denaturation has many ways, and the heat treatment method is the most common pretreatment method due to the characteristics of simple method, low investment and difficult recovery after protein denaturation.

Determination of the pretreatment temperature: treating 100ml of 10% squid protein suspension at different temperatures for 20min, adding papain, maintaining the pH of enzymolysis environment at 7.0, adding 50U/g enzyme, and allowing enzymolysis for 3 hr. The results are shown in FIG. 1.

It can be seen from FIG. 1 that the degree of hydrolysis tends to increase at the pretreatment temperature of 50-90 ℃ and reaches a maximum at 90 ℃ while the degree of hydrolysis decreases rapidly thereafter. This is because the protein is denatured by heating, weak bonds for maintaining the conformation of the protein are broken, and the nonpolar group inside the original molecule is exposed on the surface of the molecule, so that the contact sites of the protease are increased, and the hydrolysis speed is accelerated. When the temperature exceeds 90 ℃, protein molecules excessively agglutinate and polymerize to cause the compact structure of the protein, which is not beneficial to the combination of the contact sites of the enzyme and the protein, and the hydrolysis degree is reduced. Therefore, the pretreatment temperature was determined to be 90 ℃.

Determination of the preprocessing time: taking 100ml of 10% sleeve-fish protein suspension, treating at 90 deg.C for different time, adding papain, keeping pH7.0 in enzymolysis environment, adding 500U/g enzyme, and allowing enzymolysis for 3 hr. The final experimental results are shown in fig. 2.

As can be seen from FIG. 2, the maximum hydrolysis degree was reached when the temperature was 90 ℃ and the hydrolysis time was maintained for 15min, whereas the hydrolysis degree decreased after 15 min. This shows a certain trend with the change of heating temperature and time. The reason for this is that if the temperature is too high or the time is too long, the protein is excessively polymerized after thermal denaturation, and the degree of hydrolysis is reduced. Thus, the pretreatment temperature was 90 ℃ for 15 min.

Determination of technological parameters of papain and flavor enzyme for squid protein enzymolysis

Influence of hydrolysis time on squid protein enzymolysis:

the enzymolysis conditions of the papain and the flavor enzyme squid protein are as follows: the temperature is 50 ℃, the pH is 7.0, the substrate concentration is 5% (w/w), the enzyme adding amount is 500U/g of the squid, the hydrolysis time is 8 hours, after sampling every 1 hour, the hydrolysis degree of the squid is measured, and the measurement result is shown in figure 3.

As can be seen from FIG. 3, the degree of hydrolysis continuously increases with the progress of the enzymatic hydrolysis, and the increase trend gradually becomes gentle after reaching a certain value. Therefore, in order to save energy and achieve the same hydrolysis effect, the hydrolysis time of papain is determined to be 4h, and the hydrolysis time of flavor enzyme enzymolysis squid protein is determined to be 2 h.

Influence of pH on enzymolysis of squid protein

The enzymolysis conditions of the papain and the flavor enzyme squid protein are as follows: the temperature is 50 ℃, the enzyme adding amount is 500U/g squid, the substrate concentration is 5% (w/w), papain and flavor enzyme are added to keep the enzymolysis time at 4h and 2h respectively, and finally the measured hydrolysis degree is shown in figure 4.

The influence of pH on an enzymatic reaction includes two aspects, namely, the stability of the enzyme and the ability of the enzyme to catalyze the conversion of a substrate to a product. Under different pH values, the hydrolysis degree of the squid protein by the protease is different, when the hydrolysis degree is increased to a certain level, the stability of the enzyme is reduced, the enzyme activity is reduced, and the hydrolysis degree is reduced accordingly. As shown in FIG. 4, the abilities of papain and flavor enzyme to hydrolyze squid protein tend to increase and decrease with increasing pH, and the hydrolysis degree of the flavor enzyme reaches a maximum value near pH6.0 at pH6.5 of papain.

Effect of substrate concentration on enzymatic hydrolysis of Fish proteins

The enzymolysis conditions of the papain and the flavor enzyme squid protein are as follows: the temperature is 50 ℃, and the enzyme adding amount is 500U/g of squid. Then papain and flavor enzyme were added, respectively, the pH was adjusted to 6.5 and 6.0, respectively, the hydrolysis time was 4h and 2h, respectively, and then the degree of hydrolysis of the squid was measured as shown in fig. 5.

The substrate concentration has an important effect on the rate of the enzymatic reaction. The raw material is added with little water, the hydrolysis is not thorough, and the hydrolysis degree of the protein is low; the addition of more water is beneficial to the hydrolysis reaction, the hydrolysis degree is improved, but the concentration of the substrate is low, and the energy consumption generated during concentration is large. As shown in fig. 5, the smaller substrate concentration is beneficial to the enzymolysis process, because the enzymolysis reaction is performed in the aqueous solution, the presence of a large amount of water is beneficial to the diffusion of molecules, and the higher the substrate concentration is, the lower the effective concentration of water is, and the speed of the enzymolysis reaction is reduced. And (3) combining the actual production environment, considering the subsequent production process, and determining that the substrate concentrations of the papain and the flavor enzyme for hydrolyzing the squid protein are both 8% under the condition of ensuring high hydrolysis degree.

Influence of enzyme addition on degree of hydrolysis of fish protein

Maintaining the enzymolysis conditions of the papain and the flavor enzyme: the temperature was 50 ℃ and the substrate concentration was 8% (w/w), after which papain and flavourzyme were added and the pH was adjusted to 6.5 and 6.0, respectively, for hydrolysis for 4h, 2h, respectively, the results of which are shown in FIG. 6.

The amount of enzyme added can also be expressed in terms of the enzyme substrate ratio, i.e., the ratio of enzyme concentration to substrate concentration is abbreviated as (E/S). The enzyme base ratio can better reflect the reaction speed characteristic in the enzymolysis process than the enzyme concentration. When the substrate concentration is constant and the substrate concentration is not saturated by the increased amount of enzyme, the larger the E/S, the higher the reaction rate. However, the larger the E/S, the higher the production cost. Meanwhile, the larger the enzyme adding amount is, the enzyme self-inhibition may be caused, and the enzyme activity is reduced. The enzyme adding amount is taken as a measurement index, and the enzyme adding amount is substantially the same as the enzyme base ratio, so that the enzymolysis process can be clearly described. As shown in FIG. 6, the degree of hydrolysis increased with the increase of the enzyme addition amount, and the degree of hydrolysis tended to be mild when the enzyme addition amount reached a certain level, and the optimum addition amounts of papain and flavourzyme were determined to be 600U/g and 450U/g in consideration of the production cost.

Orthogonal optimization experiment of papain enzymolysis parameters

After 4h of hydrolysis time, the degree of hydrolysis of the squid protein increases slowly. The hydrolysis time was determined to be 4 hours, taking into account the effect on energy consumption and production cycle. As can be seen from the single-factor experiment of the substrate concentration, the lower the substrate concentration is, the enzymolysis process is facilitated to be carried out, and the substrate concentration is determined to be 8% by comprehensive consideration. In addition to hydrolysis time, other factors such as temperature, pH, enzyme addition, etc. have different effects on the degree of hydrolysis, so an orthogonal table (see Table 3; Table 4) was designed to determine the optimal enzymatic conditions for papain.

TABLE 3 factors and levels

TABLE 4 ANOVA TABLE

The hydrolysis degree reaches 27.5 percent.

Orthogonal experimental optimization of flavor enzyme enzymolysis parameters

According to the results of the one-factor test, the flavor enzyme hydrolysis time has less influence on the enzymatic reaction, so that it is not taken into consideration. Aiming at the different influences of other four factors on the hydrolysis degree, the enzymolysis conditions are further optimized, the designed orthogonal test table is shown in table 5, and other enzymolysis conditions are controlled: the hydrolysis time was kept for 2 h.

TABLE 5 factors and levels

TABLE 6 ANOVA TABLE

As can be seen from tables 5 and 6: the main and secondary factors influencing the degree of hydrolysis are as follows through range analysis: pH > temperature > substrate concentration > enzyme addition; it was found by further analysis of variance that pH had a significant effect on the degree of hydrolysis (p <0.05), whereas temperature, substrate concentration and enzyme addition had no significant effect on the degree of hydrolysis. Finally, the optimal enzymolysis process combination is obtained through orthogonal experiments as follows: the temperature is 45 ℃, the pH value is 6.0, the substrate concentration is 8 percent, and the enzyme adding amount is 500U/g squid. Proved by verification experiments, the hydrolysis degree of the squid hydrolysate reaches 28.1 percent.

Conclusion

The squid leftovers are used as raw materials, the proteins are denatured by adopting a heat treatment mode, exogenous protease (papain, flavor enzyme, trypsin, neutral protease and pepsin) is added for carrying out enzymolysis on the squid proteins, the hydrolysis degree is used as an index, the five proteases are comprehensively compared, and the papain and the flavor enzyme are determined to be used as the protease in the subsequent enzymolysis process.

Aiming at papain and flavor enzyme, the influence of temperature, pH, substrate concentration, hydrolysis time and enzyme addition amount on squid proteolysis is researched, and the enzymolysis conditions of the enzymes are optimized through orthogonal experiments, so that the method is finally obtained: the optimal conditions of the papain are as follows: the temperature is 50 ℃, the pH value is 7.0, and the enzyme adding amount is 600U/g squid. Through verification experiments, the hydrolysis degree reaches 29.5%. Optimal conditions for flavourzyme: the temperature is 45 ℃, the pH value is 6.0, the substrate concentration is 8%, the enzyme adding amount is 500U/g squid, and the hydrolysis degree reaches 28.1% through verification experiments.

The solubility difference of the squid leftovers divides the squid leftovers into 3 components of water-soluble protein, salt-soluble protein and insoluble protein. Freeze drying to obtain water soluble, salt soluble and insoluble protein powder with protein content of 63.67%, 48.39% and 62.84%. The protein content of the salt-soluble protein after desalting treatment can reach 82.15%, and the contents of flavor amino acid and branched chain amino acid are higher. In summary, in the expanded production test, the papain and the flavor enzyme are mainly used for enzymolysis comparison, and the flavor enzyme is more suitable for production and processing of the squid protein powder in view of economic cost.

Research on key technology of spray drying of squid leftover enzymolysis protein

The Loligo chinensis Gray has the advantages of high protein, low fat, and low calorie. The squid is rich in various amino acids necessary for human body, and the composition of the necessary amino acids is close to that of holoprotein, so that the squid is a nutritional health-care aquatic product resource with good flavor.

The method is characterized in that the squid generates about 15 to 20 percent of waste such as viscera, epidermis and the like in the process of processing the squid into shredded squid, squid rings, squid flowers or seasoning squid products, and the hydrolyzed squid protein powder is prepared by taking the waste from processing the squid leftovers as raw materials, performing enzymolysis treatment by protease and then adopting a spray drying process. The spray drying is suitable for drying heat-sensitive enzymolysis concentrated solution, and has the advantages of high drying speed, good product quality and high purity. Can save the procedures of concentration, filtration, pulverization and the like to obtain a powdery product of 30-500um, and has the advantages of short time consumption, high production efficiency and good product fluidity and instant solubility.

And dispersing the enzymolysis concentrated solution into mist-like liquid drops through an atomizer in the spray drying process, and performing heat exchange under the action of dry air to quickly evaporate a solvent in the mist-like liquid drops to obtain a powdery or granular product. Firstly, selecting three factors of air inlet temperature, feed concentration and drying aid addition amount to carry out single-factor experiment, and determining the optimal experiment process of the three factors according to the wall sticking condition in the spraying process and the quality of the hydrolyzed protein powder. And performing orthogonal experiment on the basis to determine the optimal process condition of spray drying to obtain hydrolyzed squid protein powder with better quality.

Determination of spray drying inlet air temperature

In the technological parameters of spray drying, the liquid is dispersed into mist drops by an atomizer, and heat exchange is carried out under the action of dry air to quickly evaporate the solvent in the mist drops, so as to obtain a powdery or granular product. In this spray drying process, the temperature of the incoming air is the primary factor, which directly affects the spray drying process and the quality of the final product.

And selecting different air inlet temperatures to spray-dry the squid hydrolyzed protein powder, and comparing the wall sticking condition, the water content, the amino acid nitrogen content, the color and the sensory quality of the obtained squid hydrolyzed protein powder to finally obtain the temperature which is most suitable for spray-drying. The squid hydrolyzed protein powder sensory evaluation was carried out, and the criteria are shown in the following table 7.

TABLE 7 hydrolyzed protein powder score criteria

Influence of the inlet air temperature on the physicochemical properties of the hydrolyzed protein powder: under the condition of controlling the same air inlet quantity, the higher the temperature of the drying chamber is, the earlier the drying enters the drying speed reduction stage, and the less the water content of the corresponding dried product is, so that the wall sticking phenomenon basically cannot occur; however, excessive high temperature easily causes the squid protein powder to have caramelization reaction and Maillard reaction, thereby generating burnt flavor and protein denaturation. When the temperature of the inlet air is lower, the drying speed is reduced, and the protein powder is easy to form large particles. According to the experimental results in table 8, it can be found that the hydrolyzed protein powder is less sticky to the wall when the inlet air temperature is 190 ℃, the squid taste is stronger, and the drying effect is most ideal. But the water content of the hydrolyzed squid protein powder is obviously reduced (p is less than 0.05) along with the increase of the temperature.

TABLE 8 influence of inlet air temperature on spray drying wall-sticking situation

This is due to the abundance of proteins and reducing sugars in squid, which causes the reducing sugars to undergo maillard reactions with proteins during spray drying, resulting in the loss of some amino acids, notably L-lysine. Therefore, free amino acid is lost to varying degrees as the temperature of the intake air is affected. The results in FIG. 7 show that the content of amino acid nitrogen tends to decrease as the intake air temperature continues to increase. It can be seen that the difference between the amino acid nitrogen content at the inlet air temperature of 170 ℃ and 190 ℃ is not obvious (p is more than 0.05); the hydrolyzed protein powder obtained at 150 deg.C has the lowest content of amino acid nitrogen, and the highest content at 170 deg.C.

Influence of the inlet air temperature on the sensory properties of the hydrolyzed protein powder: as can be seen from the following Table 9, the color of the squid hydrolyzed protein powder is not significantly different when the inlet air temperature is kept at 170-210 ℃ (p > 0.05). Under the condition that the temperature of the inlet air is 150 ℃, the brightness of the squid protein powder is reduced, and the red degree is increased, which is probably caused by that fog drops are not dried fully, and the water content of the protein powder is more. As can be seen from the total color difference esab, 190 ℃ is the smallest, indicating the lightest overall color. Therefore, the air inlet temperature of the protein powder in the spray drying production should not be lower than 170 ℃.

TABLE 9 Effect of different intake air temperatures on color

TABLE 10 influence of different intake air temperatures on sensory quality

As can be seen from table 10 above, the difference in sensory score at different temperatures was not particularly significant, but the resulting egg white powder score was the highest at 190 ℃ and the lowest at 150 ℃. Therefore, the optimal experimental level of the selected inlet air temperature of 190 ℃ is finally determined by comprehensively considering the influence results of the color and luster and the scores of the sensory evaluation in the table 9.

Determination of spray-dried feed concentration

The feed concentration of the feed liquid not only affects the spray drying effect, but also affects the economic cost of the spray drying. In contrast, the thermal efficiency of spray drying is low, only about 30% -40%, so that the cost can be greatly reduced by properly concentrating the feed liquid before feeding. In addition, for the same material, the proper concentration is beneficial to forming large particle size of dried particles during spray drying, and the larger particle size is beneficial to improving the fluidity and the dispersibility of the squid protein powder after spray drying and reducing the agglomeration phenomenon of the protein powder.

Influence of feed concentration on physicochemical properties of hydrolyzed protein powder: the concentration of the fed material is too low, the proportion of water in fog drops is large, the water cannot be fully evaporated in the spray drying process, and the phenomenon of wall sticking can occur; when pan feeding concentration increases, the water content of single droplet diminishes, and the drying can be more thorough under the same dry environment. As shown in Table 11, the wall sticking phenomenon was small at the feed concentration of 15% and 20%, while the wall sticking was serious at 5% and 10%, particularly at 5%. However, when the concentration is higher, the fog drops are dried from outside to inside, and because the hydrolyzed protein powder contains a certain amount of carbohydrate, a layer of compact-structure vitreous body is formed on the surface of larger liquid drops firstly, so that the migration of internal moisture is prevented, and the drying speed is reduced.

TABLE 11 Effect of feed concentration on spray-drying wall-sticking

As shown in FIG. 8, the water content of the hydrolyzed protein powder decreased with the increase of the concentration of the feed. When the concentration is increased to 15%, the water content of the hydrolyzed protein powder is the lowest and is 2.9%; however, when the concentration is increased to 20%, the water content rises back up. However, if the material is at a high temperature for a long time, protein denaturation and other phenomena easily occur to the protein in the hydrolyzed protein powder. When the concentration reaches more than 20%, not only the water content of the hydrolyzed protein powder begins to rise, but also the nutrients begin to be lost. Along with the increase of the feed concentration, the content of amino acid nitrogen contained in the hydrolyzed protein powder has a regular rising trend. This shows that under the same temperature condition, the higher the feed concentration is, the higher the content of amino acid nitrogen in the hydrolyzed protein powder is, wherein the difference between the content of amino acid nitrogen at the feed concentration of 15% and 20% is not obvious (p < 0.05).

Influence of feed concentration on sensory properties of hydrolyzed protein powder: as shown in Table 12 below, the hydrolyzed protein powder with 15% feed concentration had the highest brightness, the best color, and the smallest total color difference. Table 13 shows that the hydrolyzed protein powder has the highest sensory score and good color, flavor, taste and texture when the feed concentration is 15%. Therefore, through the analysis of data obtained by combining the two tables, 15% of the powder is ideal feeding concentration, the spraying effect is good, the water content is moderate, and the powder also has good dispersibility and fluidity.

TABLE 12 Effect of different feed concentrations on color

TABLE 13 Effect of different feed concentrations on organoleptic qualities

Determination of the amount of drying aid to be added in spray drying

The squid enzymolysis liquid contains a large amount of water and more substances such as amino acid, polypeptide, saccharide and the like, if the wall sticking phenomenon can be caused by directly carrying out spray drying, the powder yield can be greatly reduced, and the spraying effect is influenced. The softening point of the enzymolysis concentrated solution is lower than the air inlet temperature of spray drying, which is the main reason for the wall sticking phenomenon, and the wall sticking phenomenon can be eliminated by adding auxiliary materials to improve the softening point of the concentrated solution. And selecting different proportions of squid solid content and maltodextrin to carry out spray drying.

Influence of the addition amount of the drying assistant on the physicochemical property of the hydrolyzed protein powder: under the condition that amino acid, polypeptide and reducing sugar contained in the hydrolyzed protein powder coexist, browning reaction is easily caused by high-temperature treatment of the hydrolyzed protein powder, so that loss of nutrient substances is caused, and maltodextrin can play a role of drying a carrier, so that the browning reaction can be effectively inhibited, and the loss of the nutrient substances is reduced to a certain extent. As shown in FIG. 9, the water content of the hydrolyzed protein powder tended to decrease with the increase of the amount of maltodextrin added. When the ratio of the squid solid matter to the maltodextrin is 4:1, 3:1 and 2:1, the water content of the hydrolyzed protein powder is continuously reduced, and the change is obvious (p is more than 0.05). When the powder adding amount is 4:1, the content of amino acid nitrogen in the hydrolyzed protein powder is higher than that in the cases of 3:1 and 2:1 (p is less than 0.05). Therefore, in order to ensure a better spraying effect and good quality of the hydrolyzed protein powder, the powder adding amount ratio of 4:1 is determined as the optimal experimental level.

Influence of the addition amount of the drying assistant on the sensory properties of the hydrolyzed protein powder:

the maltodextrin is prepared from starch serving as a raw material, has good water solubility, can form uniform droplets in a spraying process, and has a good embedding effect and improved product flavor. As shown in Table 14, the addition of maltodextrin hardly caused the wall-sticking phenomenon, and the dispersibility and flowability of the hydrolyzed protein powder were also improved. When the addition amount is 4:1, the wall sticking is less, the dryness of the hydrolyzed protein powder is high, the color is good, and the squid taste is fragrant and gloomy.

TABLE 14 Effect of maltodextrin on spray drying

Optimizing spray drying process parameters: in order to optimize the spray drying process of the hydrolyzed squid protein powder, L is carried out on the basis of the single-factor experimental results of the three influencing factors₉The results of the orthogonal experiments are shown in tables 15 and 16.

TABLE 15 spray drying Quadrature Experimental results

As can be seen from the results of the range analysis in Table 16, the influence of the factors on the content of amino acid nitrogen in the hydrolyzed protein powder is in the following order: the powder adding amount is larger than the air inlet temperature and the material feeding concentration; the squid protein powder sensory evaluation influence is sequentially in the order of magnitude: the air inlet temperature > the powder adding amount > the feeding concentration.

TABLE 16 very poor analysis of spray-drying

The range analysis is in A₂B₁C₃Under the condition, the obtained squid powder has higher content of amino acid nitrogen. On visual analysis, the best combination of sensory analysis is A₂B₃C₃I.e. experiment No. 5. The influence of various factors on the content of amino acid nitrogen in the hydrolyzed protein powder is as follows in sequence: amount of powder added>Temperature of inlet air>Feed concentration, thus B in the first two combinations₁And B₂All can be used. It can be seen from the range analysis that₂B₃C₃The obtained squid powder has higher sensory evaluation, and the influence on the sensory evaluation of the squid powder is the air inlet temperature>Amount of powder added>The feed concentration, with the smallest influence also being B, i.e. the feed concentration. Therefore, from the viewpoint of energy saving, the feed concentration was selected to be 15%.

In summary, the optimal combination obtained by the orthogonal experiment is A₂B₃C₃When the air inlet temperature is 190 ℃, the feed concentration is 15%, and the ratio of the squid solid matter to the maltodextrin is 4:1, all indexes of the squid hydrolyzed protein powder obtained under the condition are relatively better.

Conclusion

The optimal process conditions for preparing the squid hydrolyzed protein powder by adopting the spray drying process are as follows: the air inlet temperature is 190 ℃, the material inlet concentration is 15%, the ratio of the squid solid matter to the maltodextrin is 4:1, the content of amino acid nitrogen in the finished product of the hydrolyzed protein powder is 3.989%, the water content is 2.91%, and the sensory score is 94%.

The hydrolyzed protein powder has little wall sticking phenomenon in the spray drying process, and is not easy to generate caramelization reaction and Maillard reaction. The water content of the protein powder is moderate, and the powder has better dispersibility and fluidity; has good color, no browning, fragrant squid taste and good quality.

The skilled person should understand that: although the invention has been described in terms of the above specific embodiments, the inventive concept is not limited thereto and any modification applying the inventive concept is intended to be included within the scope of the patent claims.

The above description is only a preferred embodiment of the present invention, and the protection scope of the present invention is not limited to the above embodiments, and all technical solutions belonging to the idea of the present invention belong to the protection scope of the present invention. It should be noted that modifications and embellishments within the scope of the invention may occur to those skilled in the art without departing from the principle of the invention, and are considered to be within the scope of the invention.

Claims

1. The processing method for preparing the biological enzymolysis protein by using the squid leftovers is characterized by comprising the following steps of:

(6) blending: adding fishy smell removing substance, and adjusting its content;

(7) homogenizing: homogenizing the mixed liquid;

2. The processing method for preparing the biological enzymolysis protein from the squid leftovers according to the claim 1, characterized in that when the exogenous protease for hydrolyzing the squid protein is papain, the hydrolysis temperature is 50 ℃, and the pH value of the reaction solution is adjusted to be 7.0.

3. The processing method for preparing the biological enzymolysis protein from the squid leftovers according to the claim 1, characterized in that when the exogenous protease for hydrolyzing the squid protein is the flavourzyme, the hydrolysis temperature is 45 ℃, and the pH value of the reaction solution is adjusted to 6.0.

4. The processing method for preparing the biological enzymolysis protein by the squid leftovers according to the claim 1, characterized in that the air inlet temperature during the spray drying is 190 ℃.

5. The processing method for preparing the biological enzymolysis protein by the squid leftovers according to the claim 1, characterized in that the feed concentration of the feed liquid during the spray drying is 15%.

6. The processing method for preparing the biological enzymolysis protein by using the squid leftovers according to the claim 1, characterized in that the ratio of the squid solid matters and the maltodextrin which are spray drying auxiliary agents during spray drying is 4: 1.

7. The processing method for preparing the biological enzymolysis protein by the squid leftovers according to the claim 1, characterized in that the processing temperature of the enzymolysis pretreatment is 90 ℃.