Enzyme preparation for catalyzing color enhancement and quality improvement of fruits and application of enzyme preparation

Abstract

The invention discloses an enzyme preparation for catalyzing fruit color enhancement and quality improvement and application of the enzyme preparation, and relates to the technical field of biological enzyme preparations and fruit and vegetable color enhancement and quality improvement. The enzyme preparation is a catalytic enzyme preparation which is prepared from a strain CH18 through a specific fermentation process, contains three ways of anthocyanin synthesis, has broad spectrum, and can be used for preparing a high-quality enzyme preparation for fruit color enhancement and quality improvement. The color enhancing agent has a color enhancing effect on red fruits, vegetables and color leaves. The fruit coloring and reddening agent is sprayed on the surfaces or leaf surfaces of fruits for 1-3 times, can catalyze the coloring and reddening of the fruits, and can be used for coloring fruits such as apples, red dates, winter dates, tautumn sweet persimmons, grapes, cherries, haws, tomatoes, peaches and the like and reddening colored leaves such as maple leaves and the like. The enzyme preparation belongs to a biological product, does not contain harmful substances, and ensures the quality improvement and safety of agricultural products.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological enzyme preparation and fruit color enhancement and quality improvement, and more particularly to an enzyme preparation for catalyzing fruit color enhancement and quality improvement and application thereof. BACKGROUND

[0002] With the improvement of people's living standards, the demand for fruits is growing, especially the demand for high-quality fruits is increasing. The appearance color and sugar content of fruits are important quality indicators of many fruits such as apples, peaches, grapes, tomatoes, dates, etc., and are also important manifestations of fruit commodity quality, which directly affects the market positioning and commodity price. Fruits with high sugar content, good color and rich flavor are favored by consumers and have high market value. At the same time, color is also an important commodity indicator for color plants (such as ornamental trees). The color of fruits or plants is generally affected by factors such as light, temperature, water, sugar and mineral elements. In order to improve the color of colored fruits and vegetables, measures such as increasing the light intensity of fruits and vegetables, enhancing photosynthesis, reasonable fertilization, spraying chemical preparations on the surface of fruits and vegetables, reasonable water supply, leaf picking, fruit turning, bagging, laying reflective film, and protecting the shallow root system of fruit trees can be taken.

[0003] In addition, in order to improve the color of fruits and vegetables, plant growth regulators such as prohexadione calcium, forchlorfenuron, sodium nitrophenolate, gibberellin, daminozide, paclobutrazol, naphthalene acetic acid, ethylene, and abscisic acid can be sprayed. However, such colorants usually have some side effects, which often cause the early arrival of the respiration peak of fruits, cause fruit softening, reduce their storage resistance, and shorten the shelf life. Therefore, it is an urgent problem for those skilled in the art to develop green and safe biological colorants and sugar enhancers. SUMMARY

[0004] Therefore, the present application provides an enzyme preparation for catalyzing fruit color enhancement and quality improvement and application thereof.

[0005] In order to achieve the above purpose, the present application adopts the following technical solutions:

[0006] An enzyme preparation for catalyzing fruit color enhancement and quality improvement is prepared by the following method:

[0007] The strain CH18 is activated and cultured to obtain a seed liquid, which is inoculated into a potato liquid fermentation medium (PDA) or a potato powder immersion fermentation medium (PDAE) at an inoculation amount of 1%-5% to be cultured, wherein a carbon source is one or a combination of glucose, sucrose, maltose and starch, and the addition amount is 20 g / L-40 g / L. The fermentation temperature is 20°C-30°C, and the rotation speed is 160 r / min-200 r / min. The fermentation is carried out for 2-5 days, and the specific harvesting time is determined according to the change trend of the solid content or the protein content. The fermentation liquid obtained by harvesting is broken, and the supernatant is obtained. Nutrient elements, a surfactant and amino acids are added and mixed to obtain an enzyme preparation.

[0008] As a preferred technical solution, the basic components of the potato liquid fermentation medium are fresh peeled potato 200-250 g, one or a combination of glucose, sucrose, maltose and starch, and the addition amount is 20 g / L-40 g / L, and water 1000 ml. The basic components of the potato powder immersion fermentation medium are powder immersion 5-8 g, one or a combination of glucose, sucrose, maltose and starch, and the addition amount is 20 g / L-40 g / L, and water 1000 ml.

[0009] As a preferred technical solution, the preparation method of the seed liquid is that the strain CH18 stored in a test tube slant or an EP tube is inoculated into a potato liquid medium, and activated and cultured at a temperature of 20°C-30°C and a rotation speed of 150 r / min-200 r / min for 2-6 days. The seed liquid is obtained by further transferring and activating the culture for 1-2 generations.

[0010] As a preferred technical solution, the enzyme preparation can be prepared by using the broken fermentation stock solution (referred to as PD obtained by PDA fermentation and AE obtained by PDAE fermentation) or the dilution liquid (referred to as PD# obtained by dilution of PD and AE# obtained by dilution of AE) diluted by 1-20 times.

[0011] As a preferred technical solution, the enzyme preparation contains a complete enzyme system for anthocyanin synthesis, that is, a catalytic enzyme system in the three pathways of anthocyanin synthesis.

[0012] As a preferred technical solution, the nutrient elements are one or a combination of potassium dihydrogen phosphate, calcium elements and boron elements, the surfactant is one or a combination of soil temperature 80 and sodium alginate, and the amino acid is phenylalanine.

[0013] More preferably, the potassium dihydrogen phosphate is added in an amount of 0.1-0.6%, the calcium element is added in an amount of 0.1-0.3%, the boron element is added in an amount of 0.01-0.20%, the surfactant soil temperature 80 is added in an amount of 0.15-0.80%, and the sodium alginate is added in an amount of 0.05-0.30%; and the phenylalanine is added in an amount of 10-100 mg / L.

[0014] As a preferred technical solution, the harvesting time is when the solid content in the fermentation broth is reduced to 25%-50% of the initial solid content, or when the inflection point of the protein content rising trend in the fermentation broth appears, the fermentation is stopped, and the fermentation broth is harvested.

[0015] As a preferred technical solution, the enzyme preparation is sprayed on the fruit surface and leaf surface 1-3 times within 5-40 days from the beginning of the fruit or color leaf color changing period, which can catalyze the fruit and color leaf color changing and increasing redness, and can increase the sugar content of the fruit.

[0016] Another object of the present application is to provide the use of the above-mentioned enzyme preparation in coloring and / or increasing the sugar content of plants.

[0017] Another object of the present application is to provide the use of the above-mentioned enzyme preparation in coloring and / or increasing the sugar content of plants.

[0018] Through the above technical solution, compared with the prior art, the present application provides a biological enzyme preparation for catalyzing fruit coloring and quality improvement and its application, which has the following beneficial effects:

[0019] (1) The enzyme preparation sprayed in the present application contains a complete set of enzymes for promoting the formation of anthocyanins, i.e. contains catalytic enzyme systems of three anthocyanin synthesis pathways, has broad spectrum, and has coloring effect on all red-purple fruits and color leaf plants. By regulating the anthocyanin color substance synthesis and metabolic pathway through enzyme reaction, the synthesis and accumulation of anthocyanins in fruits are promoted, so as to increase the redness of fruits.

[0020] (2) The enzyme preparation is obtained by fermentation of the strain CH18, and can be sprayed on the fruit surface and leaf surface 1-3 times, which can catalyze the fruit and color leaf color changing and increasing redness, and can increase the sugar content of the fruit, and can be used for fruit coloring and color leaf redness increasing of apple, board jujube, Taqiu sweet persimmon, grape, cherry, winter jujube, hawthorn, tomato, peach, etc.

[0021] (3) The enzyme preparation is produced by fermentation, has low cost, and can be produced industrially.

[0022] (4) The enzyme preparation is a biological product, does not contain harmful substances, and ensures the quality and safety of agricultural products. BRIEF DESCRIPTION OF DRAWINGS

[0023] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the following will briefly introduce the drawings needed to be used in the embodiments or prior art description. Obviously, the drawings described below are only a part of the embodiments of the present application, and not all embodiments. Based on the embodiments of the present application, all other embodiments obtained by those of ordinary skill in the art without creative labor are within the scope of protection of the present application.

[0024] Figure 1 Positive ion mode chromatogram of the enzyme preparation stock solution of the present application.

[0025] Figure 2 Negative ion mode chromatogram of the enzyme preparation stock solution of the present application.

[0026] Figure 3 Chromatogram and mass spectrum of dihydroquercetin, a cyanidin synthesis substrate in the enzyme preparation stock solution of the present application;

[0027] Wherein A is a chromatogram, B is a primary mass spectrum, and C is a secondary mass spectrum.

[0028] Figure 4 Chromatogram and mass spectrum of dihydromorin, a cyanidin synthesis substrate in the enzyme preparation stock solution of the present application; wherein A is a chromatogram, B is a primary mass spectrum, and C is a secondary mass spectrum.

[0029] Figure 5 Chromatogram and mass spectrum of dihydromyricetin, a cyanidin synthesis substrate in the enzyme preparation stock solution of the present application; wherein A is a chromatogram, B is a primary mass spectrum, and C is a secondary mass spectrum.

[0030] Figure 6 Pathway of CH18 strain synthesizing cyanidin of the present application.

[0031] Figure 7 Summary of plant cyanidin synthesis pathway.

[0032] Figure 8 Gene amplification result of CH18 cyanidin synthesis enzyme. DETAILED DESCRIPTION

[0033] The technical solutions in the embodiments of the present application will be described clearly and completely below in combination with the drawings in the embodiments of the present application. Obviously, the described embodiments are only a part of the embodiments of the present application, and not all embodiments. Based on the embodiments of the present application, all other embodiments obtained by those of ordinary skill in the art without creative labor are within the scope of protection of the present application.

[0034] The basic components of the potato liquid fermentation medium (PDA) of the present application are 200-250 g of fresh peeled potatoes, one or a combination of glucose or sucrose or maltose or starch, added in an amount of 20 g / L-40 g / L, and 1000 ml of water.

[0035] The basic components of the potato powder immersion fermentation medium (PDAE) of the present application are 5-8 g of powder immersion, one or a combination of glucose or sucrose or maltose or starch, added in an amount of 20 g / L-40 g / L, and 1000 ml of water.

[0036] Example 1 Preparation of enzyme preparation

[0037] 1. Take the strain CH18 (which has been published in CN108823101B, with the preservation number of CGMCC NO. 14784) stored in a test tube slant or EP tube, inoculate it in a potato liquid fermentation medium (PDA), activate and culture for 4 days at a temperature of 28°C and a rotation speed of 180 r / min, continue to transfer and culture for 1-2 generations to obtain a seed liquid.

[0038] 2. Then inoculate the seed liquid into the potato liquid fermentation medium (PDA) or the potato powder immersion medium (PDAE) according to a 3% inoculation amount. The fermentation temperature is 25°C, the rotation speed is 180 r / min, and the fermentation is carried out for 3 days. When the solid content in the fermentation liquid is reduced to 25%-50% of the initial solid content, or the inflection point of the upward trend of the protein content appears, stop the fermentation and collect the fermentation liquid.

[0039] 3. Crush the fermentation liquid collected above, take the supernatant, add nutritional elements, surfactants and amino acids, and mix well to obtain an enzyme preparation. The enzyme preparation obtained above contains a complete set of enzymes for anthocyanin synthesis, i.e. catalytic enzymes in the three pathways of anthocyanin synthesis, nutritional elements such as potassium dihydrogen phosphate, calcium and boron; surfactants such as Tween 80 or sodium alginate, and amino acids such as phenylalanine. The added nutritional elements such as potassium dihydrogen phosphate are added in an amount of 0.35%, the added calcium is added in an amount of 0.2%, the added boron is added in an amount of 0.15%; the added surfactants such as Tween 80 are added in an amount of 0.3%, the added sodium alginate is added in an amount of 0.15%; and the added amino acids such as phenylalanine are added in an amount of 30 mg / L.

[0040] When preparing the enzyme preparation, the crushed fermentation stock solution (referred to as PD obtained by PDA fermentation and AE obtained by PDAE fermentation) or the diluted solution diluted by 1-20 times (referred to as PD# obtained by PD dilution and AE# obtained by AE fermentation) can be used for configuration and use.

[0041] Example 2 Preparation of enzyme preparation

[0042] 1. Take the test tube slope or EP tube preserved strain CH18 (the strain has been published in CN108823101B, and the preservation number is CGMCC NO.14784), inoculate in potato liquid fermentation medium (PDA), activate culture at 20℃, 200r / min for 6d, continue to transfer culture for 1-2 generations, and obtain seed liquid.

[0043] 2. Then inoculate the seed liquid into potato liquid fermentation medium (PDA) or potato powder immersion medium (PDAE) according to 5% inoculation amount. The fermentation temperature is 20℃, the rotation speed is 200r / min, and the fermentation is carried out for 5d. When the solid content in the fermentation liquid is reduced to 25%-50% of the initial solid content, or the inflection point of the rising trend of protein content appears, stop the fermentation, and collect the fermentation liquid.

[0044] 3. The fermentation liquid collected above is broken, the supernatant is taken, and nutritional elements, surfactants and amino acids are added to obtain enzyme preparation. The enzyme preparation obtained above contains a complete set of enzymes for anthocyanin synthesis, i.e. catalytic enzyme system in the three pathways of anthocyanin synthesis, nutritional elements such as potassium dihydrogen phosphate, calcium and boron; surfactants such as tween 80 or sodium alginate, and amino acid is phenylalanine. The added nutritional elements such as potassium dihydrogen phosphate are 0.1%, the added calcium is 0.3%, the added boron is 0.20%; the added surfactants such as tween 80 are 0.15%, the added sodium alginate is 0.30%; and the added amino acid such as phenylalanine is 10mg / L.

[0045] When preparing the enzyme preparation, the broken fermentation stock solution (referred to as PD obtained by PDA fermentation, and referred to as AE obtained by PDAE fermentation) can be used, or the dilution liquid diluted to 1-20 (referred to as PD# obtained by PD dilution, and referred to as AE# obtained by AE fermentation) can be used.

[0046] Example 3 Preparation of enzyme preparation

[0047] 1. Take the test tube slope or EP tube preserved strain CH18 (the strain has been published in CN108823101B, and the preservation number is CGMCC NO.14784), inoculate in potato liquid fermentation medium (PDA), activate culture at 20℃, 200r / min for 6d, continue to transfer culture for 1-2 generations, and obtain seed liquid.

[0048] 2. Then, inoculate the seed liquid into potato liquid fermentation medium (PDA) or potato powder immersion medium (PDAE) according to 1% inoculation amount. The fermentation temperature is 30°C, and the rotation speed is 160 r / min. Ferment for 2 days. When the solid content in the fermentation liquid is reduced to 25%-50% of the initial solid content, or the inflection point of the rising trend of the protein content appears, stop the fermentation, and collect the fermentation liquid.

[0049] 3. Crush the fermentation liquid collected above, take the supernatant, add nutritional elements, surfactants, and amino acids, and mix to obtain an enzyme preparation. The enzyme preparation obtained above contains a complete set of enzymes for anthocyanin synthesis, i.e. catalytic enzymes in the three pathways of anthocyanin synthesis, nutritional elements such as potassium dihydrogen phosphate, calcium, and boron; surfactants such as Tween 80 or sodium alginate, and amino acids such as phenylalanine. The added amount of the nutritional elements such as potassium dihydrogen phosphate is 0.6%, the added amount of calcium is 0.1%, and the added amount of boron is 0.01%; the added amount of the surfactants such as Tween 80 is 0.80%, the added amount of sodium alginate is 0.05%, and the added amount of the amino acid such as phenylalanine is 100 mg / L.

[0050] When preparing the enzyme preparation, the crushed fermentation stock solution (referred to as PD obtained by PDA fermentation, and referred to as AE obtained by PDAE fermentation) can be used, or the diluted solution diluted by 1-20 times (referred to as PD# obtained by PD dilution, and referred to as AE# obtained by AE fermentation) can be used.

[0051] Example 4 Characterization of the enzyme preparation

[0052] Based on the determination results of liquid chromatography-mass spectrometry, the intermediate products of anthocyanin synthesis by the strain CH18 were analyzed and characterized, and the key enzyme genes in the metabolic pathway of CH18 were cloned and characterized.

[0053] 1. The intermediate products of the fermentation of the strain CH18 obtained in Example 1 were analyzed by high performance liquid chromatography-mass spectrometry, and a plurality of components of the intermediate products were obtained.

[0054] (1) Fermentation liquid sample treatment

[0055] Concentrate 0.8 mL of the sample by rotary evaporation, add 200 μL of 80% methanol for redissolution, and vortex for 1 min; ultrasonic at 4°C for 30 min;

[0056] Stand at -20°C for 1 hour;

[0057] Centrifuge at 4°C at 12000 rpm for 15 min;

[0058] Transfer 200 μL of the supernatant, add 5 μL of an internal standard (1 mg / mL dichlorophenylalanine), mix, and then transfer into an injection vial.

[0059] (2) On-machine detection

[0060] Instrumental analysis platform: LC-MS (Waters, UPLC; Thermo, Q Exactive)

[0061] Column: ACQUITY UPLC HSS T3 (2.1*100mm 1.8μm)

[0062] Chromatographic separation conditions: Column temperature 40℃; flow rate 0.3mL / min; mobile phase composition A: water + 0.05% formic acid, B: acetonitrile; sample size: 5μL, autosampler temperature 4℃.

[0063] Elution program: see Table 1.

[0064] Table 1 Mobile phase gradient elution program

[0065]

[0066] (3) Mass spectrometry detection parameters

[0067] Positive mode: heater temperature 300℃; sheath gas flow rate: 45arb; auxiliary gas flow rate: 15arb; tail gas flow rate: 1arb; electrospray voltage: 3.0KV; capillary temperature: 350℃; S-Lens RF Level, 30%.

[0068] Negative mode: heater temperature 300℃; sheath gas flow rate: 45arb; auxiliary gas flow rate: 15arb; tail gas flow rate: 1arb; electrospray voltage: 3.2KV; capillary temperature: 350℃; S-Lens RF Level, 60%.

[0069] Scan mode: primary full scan (Full Scan, m / z 70~1050) and data-dependent secondary mass spectrometry scan (dd-MS2, TopN=10); resolution: 70,000 (primary mass spectrum) & 17,500 (secondary mass spectrum). Collision mode: high-energy collision-induced dissociation (HCD). See Figures 1 and 2 for positive ion mode chromatogram and negative ion mode chromatogram results. Figure 1 and Figures 3 and 4. Figure 2 .

[0070] (4) Analysis of intermediate products synthesized by strain CH18

[0071] According to the results of liquid chromatography-mass spectrometry analysis, strain CH18 produced a variety of intermediate products during fermentation.

[0072] The data of molecular mass, chromatogram, mass spectrum, etc. of all substances were derived from liquid chromatography-mass spectrometry determination. According to the results of primary mass spectrum and secondary mass spectrum and m / z, the specific intermediate product types were resolved.

[0073] By analyzing the CH18 metabolic intermediates, a total of 14 molecular weight matching intermediates were obtained, which were phenylalanine, p-coumaric acid, chalcone, naringenin, dihydroquercetin, dihydromorin, dihydromyricetin, procyanidin A2, paeonol, prodelphinidin, leuco delphinidin, leucogenidin, paeoniflorin 3-rhamnoside 5-glucoside and pelargonidin 3-acetyl 5-glucoside. Among them, dihydroquercetin (see attached Figure 3 ), dihydromorin (see attached Figure 4 ) and dihydromyricetin (see attached Figure 5 ) are three substrates of anthocyanin metabolic pathways, and the different substrates result in different anthocyanin metabolic pathways and products.

[0074] According to the types of intermediates, the anthocyanin metabolic pathway of strain CH18 is characterized (see attached Figure 6 ). It is known that there are three pathways for plants to synthesize anthocyanins from phenylalanine (see attached Figure 7 ), namely delphinidin pathway, pelargonidin pathway and cyanidin pathway, but for a single plant species, it often contains only 1-2 pathways. The present application discloses for the first time that the synthesis of anthocyanins by microorganisms (strain CH18) covers the three synthesis pathways of anthocyanins in the plant kingdom.

[0075] 2. Key enzyme gene for synthesizing anthocyanins by strain CH18

[0076] There have been a lot of researches on key enzyme genes for synthesizing anthocyanins and coupled metabolic products in plants, but there is no research report in microorganisms. The present application uses strain CH18 to mine the key enzyme genes for synthesizing anthocyanins and coupled metabolic products in microorganisms. For the first time, total RNA of CH18 is extracted, primers are designed (see Table 2), PCR cloning is performed, and four key enzyme genes for synthesizing anthocyanins of CH18 strain are cloned for the first time, and the results are shown in attached Figure 8 (M: marker; Lane 1: BitAT; Lane 2: BitMT; Lane 3: BitDFR; Lane 4: BitCHS.), which are named BitCHS, BitDFR, BitMT and BitAT in turn, and their sequence lengths are 1048bp, 1352bp, 769bp and 439bp, respectively.

[0077] Table 2: Primers designed for conserved sequences of key enzyme genes of strain CH18

[0078]

[0079] DFR is the key enzyme in the plant anthocyanin synthesis pathway that converts dihydroflavonols into anthocyanidins, and determines the formation pathway of anthocyanins. DFR has three substrates, namely dihydroquercetin, dihydrokaempferol and dihydromyricetin, and different substrates result in different anthocyanin metabolic pathways and products.

[0080] The subject group first detected three substrates associated with the BitDFR enzyme of the CH18 strain, namely dihydroquercetin, dihydrokaempferol and dihydromyricetin, and also detected the key BitDFR enzyme catalytic metabolism product leucodelphinidin, leucopelargonidin, and the product paeoniflorin 3-rhamnoside 5-glucoside.

[0081] The 134th amino acid encoded by the BitDFR of CH18 clone is lysine, and the 145th is proline, so BitDFR is not of the Asn / Asp type, and the amino acid types at these two sites are different from most plants.

[0082] The BitDFR of CH18 has three substrates, and all are detected in the products, that is, CH18 contains three anthocyanin synthesis pathways, covering the three anthocyanin synthesis pathways of plants, and has broad spectrum, which is different from the plant kingdom. For example, the DFR enzyme of 82 varieties of chrysanthemum can only catalyze dihydroquercetin, that is, only the cyanidin metabolic pathway exists; the DFR enzyme of anthurium can only catalyze dihydroquercetin and dihydroflavonols, that is, only the cyanidin and pelargonidin metabolic pathways exist; the DFR enzyme of peony cannot catalyze dihydromyricetin, that is, the delphinidin metabolic pathway does not exist. The strain CH18 of the present application covers the enzyme system of the three anthocyanin synthesis pathways of plants, which is obtained for the spectrum anthocyanin enzyme system, opens up a new path, and provides support for its broad-spectrum application.

[0083] Example 5: Color enhancement and quality improvement effect of enzyme preparation spraying

[0084] In 2023-2024, the subject group used strain CH18 to ferment (20-37℃, 150-200r / min) for 7-10 days to produce a large amount of anthocyanins, and the fermentation broth solid (sugar) content was close to zero (0.0-0.1%) of the fermentation broth. The AE and AD enzyme preparation liquids were configured, and the fruit was sprayed 1-2 times during the fruit color changing period. After spraying, 20d of continuous tracking observation was carried out, and compared with the control group without spraying, it was found that the color enhancement effect of the fruit surface of the experimental group was not obvious.

[0085] The fermentation conditions of the strain CH18 are further optimized by the present research group, the fermentation formula and scheme of Example 1 are adopted, the fermentation time is shortened, and the auxiliary materials are further added, and the biological enzyme preparation is prepared, and the spraying method is as follows: the enzyme preparation is sprayed on the fruit surface or leaf surface 1-3 times within 5d-40d from the beginning of the fruit or color leaf color changing period to the harvesting, and the spraying time interval is 7-10d.

[0086] In the present application, the fruit weight is determined by weighing method; the fruit solid content (sugar content) is determined by a SW-55D handheld fruit sugar meter after juicing, and the average value is taken; and the fruit color is calculated by the red color occupying the entire fruit surface area.

[0087] The enzyme preparation is sprayed on various plants, and the following results are obtained (see Tables 3, 4, 5, 6 and 7).

[0088] Table 3 Test results of spraying enzyme preparation on Jishan plate jujube

[0089]

[0090]

[0091] The results show that spraying the enzyme preparation of Example 1 of the present application can increase the sugar content of Jishan plate jujube by 17.29-19.3%, and can also increase the fruit color by 15-30%.

[0092] Table 4 Test results of spraying enzyme preparation on hawthorn

[0093]

[0094] The results show that spraying the enzyme preparation of Example 1 of the present application can increase the sugar content of hawthorn fruit by 2.0-7.3%, and can also significantly increase the fruit color.

[0095] Table 5 Test results of spraying enzyme preparation on apple

[0096]

[0097] The results show that spraying the enzyme preparation of Example 1 of the present application can increase the sugar content of apple by 3.33-11.61%, and can also increase the fruit color by 5%-25%.

[0098] Table 6 Test results of spraying enzyme preparation on Taqiu sweet persimmon

[0099]

[0100] The results show that spraying the enzyme preparation of Example 1 of the application can increase the sugar content of Taqiu sweet persimmon by 12.9-15.3%, and can also increase the flesh color.

[0101] Table 7 Test results of spraying enzyme preparation on winter jujube

[0102]

[0103] The results show that spraying the enzyme preparation can increase the sugar content of winter jujube by 15.61%. At the same time, it can significantly increase the redness.

[0104] In summary, the enzyme preparation of the application has obvious effects on color enhancement and sugar content increase of red fruits, and can be applied to red fruit and colorful leaf redness enhancement, fruit sugar content increase and quality improvement.

[0105] The above description of the various embodiments in the specification only demonstrates a part of experimental fruit types or colorful leaf plants, and is used to illustrate the effect of the enzyme preparation. Other fruits such as grapes and maple leaves also have effects.

[0106] The various embodiments in the specification are described in a progressive manner, and each embodiment focuses on the differences from other embodiments. The same and similar parts between the various embodiments can be referred to each other.

[0107] The above description of the disclosed embodiments enables those skilled in the art to implement or use the application. Various modifications to the embodiments will be apparent to those skilled in the art, and the general principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the application. Therefore, the application will not be limited to the embodiments shown herein, but will conform to the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. An enzyme preparation for catalyzing the color enhancement and quality improvement of fruits, characterized in that, It is prepared by the following method: The CH18 strain was activated and cultured to obtain a seed culture. This seed culture was then inoculated into potato liquid fermentation medium or potato starch fermentation medium at an inoculation rate of 1%-5%. The medium was supplemented with one or more of the following carbon sources: glucose, sucrose, maltose, and starch, at a concentration of 20-40 g / L. The fermentation temperature was 20-30℃, and the rotation speed was 160-200 r / min. Fermentation lasted 2-5 days. The specific harvesting time was determined based on changes in solid content or protein content. The harvested fermentation broth was then crushed, and the supernatant was added with nutrients, surfactants, and amino acids. After mixing, the enzyme preparation was obtained.

2. The enzyme preparation for catalyzing fruit color enhancement and quality improvement according to claim 1, characterized in that, The basic components of the potato liquid fermentation medium are 200-250g of fresh peeled potatoes, one or more of glucose, sucrose, maltose, or starch, added at a rate of 20g / L-40g / L, and 1000ml of water; the basic components of the potato extract fermentation medium are 5-8g of extract powder, one or more of glucose, sucrose, maltose, or starch, added at a rate of 20g / L-40g / L, and 1000ml of water.

3. The enzyme preparation for catalyzing fruit color enhancement and quality improvement according to claim 1, characterized in that, The seed culture is prepared as follows: strain CH18, preserved in test tube slant or EP tube, is inoculated into potato liquid culture medium and activated at 20℃-30℃ and 150r / min-200r / min for 2-6 days. After 1-2 generations of subculturing, the seed culture is obtained.

4. The enzyme preparation for catalyzing fruit color enhancement and quality improvement according to claim 1, characterized in that, When preparing the enzyme preparation, it can be prepared using crushed fermentation stock solution, or it can be prepared by diluting the crushed solution to a concentration of 1-20 times.

5. The enzyme preparation for catalyzing fruit color enhancement and quality improvement according to claim 1, characterized in that, The enzyme preparation contains a complete enzyme system for anthocyanin synthesis, namely the catalytic enzyme system in the three pathways of anthocyanin synthesis.

6. The enzyme preparation for catalyzing fruit color enhancement and quality improvement according to claim 1, characterized in that, The nutrient element is one or more of potassium dihydrogen phosphate, calcium, and boron; the surfactant is one or more of Tween 80 and sodium alginate; and the amino acid is phenylalanine.

7. The enzyme preparation for catalyzing fruit color enhancement and quality improvement according to claim 1, characterized in that, The addition amounts of potassium dihydrogen phosphate are 0.1-0.6%, calcium is 0.1-0.3%, and boron is 0.01-0.20%; the addition amounts of surfactant Tween 80 are 0.15-0.80%, sodium alginate is 0.05-0.30%; and phenylalanine is 10-100 mg / L.

8. The enzyme preparation for catalyzing fruit color enhancement and quality improvement according to claim 1, characterized in that, The harvesting time is when the solid content in the fermentation broth drops to 25%-50% of the initial solid content, or when the upward trend of the protein content in the fermentation broth reaches an inflection point, at which point fermentation is stopped and the fermentation broth is harvested.

9. The use of the enzyme preparation according to any one of claims 1-8 in enhancing the color and / or increasing the sugar content of plants.

10. The use of the enzyme preparation according to any one of claims 1-8 in the preparation of plant color-enhancing and / or sugar-increasing preparations.

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