CN115896209A - A kind of extraction method of feijoa pectin rich in RG-I high ester - Google Patents

Abstract

The invention discloses a method for extracting feijoa pectin rich in RG‑I high esters, comprising the following steps: pretreating feijoa pulp into small pieces and inactivating pectinase, then using 50mM citric acid Pectin is extracted from the sodium salt solution in a microwave oven, then cooled, enzymatically hydrolyzed, centrifuged, purified with ethanol, and dried to obtain high-ester pectin rich in RG‑I. The extraction method disclosed by the invention can fully extract the pectin in the feijoa pulp by using microwave-assisted enzymolysis, shortens the enzyme treatment time, and greatly improves the extraction efficiency and product yield; and the pectin prepared by the invention It is a high degree of esterification pectin with a more complete structure, RG-Ι content of 41.38%, and a higher viscosity.

Description

A kind of extraction method of feijoa pectin rich in RG-I high ester

technical field

The invention belongs to the technical field of pectin extraction, and in particular relates to an extraction method of feijoa pectin rich in RG-I high ester.

Background technique

Feijoa, also known as Brazilian guava, is a perennial subtropical evergreen shrub plant belonging to the genus Myrtaceae South America. It belongs to a class of fruits that have emerged in my country in recent years. Feijoa is rich in nutrients and functional active ingredients, such as protein, vitamins, minerals and polyphenols. According to reports, the pectin content of feijoa fruit has reached 20%, so feijoa fruit is a treasure house of pectin resources, and the large-scale planting of feijoa fruit in my country needs to be promoted, and its processed products need to be developed. Related processed products Especially pectin extraction reports and patents are seldom.

Pectin is an acidic heteropolysaccharide widely present in the intercellular layer of the cell wall of dicotyledonous plants. Pectin is widely used in food, medicine, cosmetics and other industries because of its good emulsification, gelling, stability and thickening properties. At the same time, pectin is also a natural water-soluble dietary fiber, which can reduce the risk of cancer, diabetes, and obesity, regulate the microenvironment of the human intestinal tract, and reduce blood lipids. High-ester pectin refers to pectin with a degree of esterification (the ratio of methyl-esterified galacturonic acid units to the total galacturonic acid units in pectin) ≥ 50%, because its morphological structure is closest to natural pectin molecules , has a good application prospect. The degree of esterification will affect the structure and functional properties of pectin and the ability to form complexes with other substances. It is important to keep the complete structure of pectin as much as possible and improve the performance of pectin for better application in food and other industries. significance.

The main structural domains of pectin include homogalacturonan (HG), rhamnogalacturonan I (RG-I), rhamnogalacturonan II (RG-II), and A small amount of substituted galacturonan. Commercial pectin is mainly composed of HG domains and is often used as a thickener and gelling agent in food-related fields. Using high-temperature strong acid can remove most of the RG-I region to improve the stability and gel properties of pectin. More and more studies have shown that RG-I-rich pectin has better anti-cancer efficacy, regulation of chronic metabolic diseases, immune activity and prebiotic properties, etc. Therefore, how to scientifically extract pectin with complete RG-I has attracted the attention of more and more researchers and consumers.

The existing pectin extraction methods mainly include ion exchange method, acid/alkali extraction method, supercritical extraction method and biological enzyme method, etc., but the effect of these extraction methods is not ideal. In addition, strong acid or strong alkali is not only corrosive to equipment, but also the extracted wastewater will cause environmental pollution. Therefore, how to prepare high-ester pectin with complete structure (RG-I), good performance and high yield is one of the problems to be solved at present.

Contents of the invention

The purpose of the present invention is to provide a method for extracting feijoa pectin rich in RG-I high ester, which can obtain feijoa high ester pectin rich in RG-I structure, good quality and high yield.

For reaching above-mentioned object, the present invention provides a kind of extraction method of the feijoa pectin that is rich in RG-1 high ester, comprises the following steps:

(1) Boiling the feijoa pulp and drying it until the water content is lower than 6%, and mixing the powder and the buffer evenly after crushing;

(2) Microwave heat treatment of the mixed mixed liquid followed by enzymatic hydrolysis, centrifugation and purification, followed by drying the pectin;

The power of the microwave heating treatment is 350-400W, and the time is 5-15min.

Further, the boiling temperature in step (1) is 85-100° C., and the boiling time is 3-8 minutes.

Further, the drying temperature in step (1) is 45-55°C.

Further, the pulverized powder in step (1) is passed through a 60-mesh sieve.

Further, the ratio of powder to buffer in step (1) is 10 g: 120-180 mL, and the buffer is 50 mM sodium citrate buffer with a pH of 6.8.

Further, the enzymatic hydrolysis process is as follows: after mixing the β-glucosidase and feijoa powder uniformly according to the amount of 250U/g, enzymolysis treatment for 90-100min, the enzymolysis temperature is 45-55°C, and the enzymolysis The speed is 120-180rpm.

Further, after the enzymatic hydrolysis in step (2), the extracted solution is heated in a water bath environment at 90-100° C. for 3-5 minutes.

Further, the centrifugation speed in step (2) is 4500-5500 rpm, and the centrifugation time is 8-12 minutes.

Further, the purification specifically includes the following steps: collect the supernatant after centrifugation, add twice the volume of absolute ethanol to the supernatant, mix well and let it settle for 2 hours, centrifuge at 4500-5500rpm after the pectin is precipitated, and use Clean the pectin in the test tube 2-3 times with absolute ethanol.

Further, the drying temperature of pectin is 45-60°C.

In summary, the present invention has the following advantages:

1. The extraction method disclosed in the present invention can fully extract the pectin in the feijoa pulp through microwave-assisted enzymolysis, shorten the enzyme treatment time, and greatly improve the extraction efficiency and product yield; and the fruit prepared by the present invention The gum is pectin with a high degree of esterification (68.43%), with a more complete structure, an RGI content of 41.38%, and a higher viscosity.

2. The extraction method disclosed by the present invention has low energy consumption and little pollution, improves the utilization value of feijoa fruit and provides theoretical guidance for further development and utilization of feijoa fruit.

3, the method of the present invention has kept the RG-I regional structure of pectin more, has extracted the high-ester pectin that is rich in RG-I, wherein can use biological enzyme again after the feijoa pulp by microwave treatment The gum is separated from the raw material, and more protopectin is converted into soluble pectin to be extracted, so the extraction method disclosed in the present invention can obtain high-yield pectin.

Description of drawings

Figure 1 shows the steady shear flow curves of pectin obtained by microwave-assisted enzymatic method (MEHE), enzymatic method (EHE) and conventional hot acid water method (CHE).

Detailed ways

The principles and features of the present invention are described below in conjunction with the examples, which are only used to explain the present invention, and are not intended to limit the scope of the present invention. Those who do not indicate the specific conditions in the examples are carried out according to the conventional conditions or the conditions suggested by the manufacturer. The reagents or instruments used were not indicated by the manufacturer, and they were all conventional products that could be purchased from the market.

The microwave heating method that adopts in the present invention is to use microwave oven heating method, and the microwave oven model that uses is MM823ESJ-PA Midea microwave oven, and the stall during heating is low fire. Therefore, the present invention also adds a detection method for the frequency of the middle and low fire gears, so as to avoid the problem of unclear heating parameters. The detection method is: use IEC705 method to detect the microwave power. First, prepare two 500mL plastic beakers and fill them with 500mL of water at a temperature of about 10°C, place the beakers in the center of the cooking area, set the microwave oven to full power and turn on the power, let the microwave oven run precisely for 60s, and the magnetic control can work Temperature 2s. The beakers were then removed from the microwave oven, and each beaker was stirred with a magnetic stirrer immediately before the temperature of each beaker was measured with a thermometer, and the temperature was measured quickly, and each group was repeated twice to obtain an average value.

Example 1

The present embodiment provides a kind of extraction method that is rich in the feijoa pectin of RG-1 high ester, comprises the following steps:

(1) The plucked feijoas were washed, dried, peeled and cut into pieces, then placed in deionized water at 90° C. and boiled for 5 minutes to inactivate pectinase. After properly controlling the water, put it in a blast drying oven at 50°C and dry it until the water content is less than 6%. After pulverizing with a pulverizer, pass through a 60-mesh sieve to make feijoa pulp powder (pulp and pulp) and seal it in a self-sealing container. bag, and store in a desiccator for later use.

(2) Take 10g of feijoa powder from step (1) in a 250mL conical flask, add 150mL of 50mM sodium citrate buffer (solid-to-liquid ratio 1:15), adjust the pH to 6.8 with citric acid, and mix well.

(3) Put the mixture of (2) into a microwave oven (MM823ESJ-PA Midea microwave oven), set the heat to medium-low heat (380W IEC) for 9 minutes, take it out and cool it until it is not hot.

(4) Weigh β-glucosidase (250U/g, the ratio of enzyme activity to substrate) in a conical flask, mix evenly, and carry out enzymolysis treatment in a constant temperature shaking incubator for 99min, and the enzymolysis temperature is set at 50°C, the rotating speed of the shaker is 150rpm.

(5) After the extraction, heat in a 95° C. water bath for 5 minutes to inactivate the enzyme.

(6) Centrifuge the extracted solution at 5000 rpm for 10 min, collect the supernatant, wash the residue twice with distilled water, and record the volume after combining the supernatant.

(7) Add twice the volume of absolute ethanol to the supernatant, mix well and let it settle for two hours, centrifuge at 5000 rpm after the pectin is precipitated, and wash the pectin in the test tube twice with absolute ethanol.

(8) Dry the cleaned pectin in an electric blast drying oven at a set temperature of 50° C., grind it with a mortar and pass it through a 100-mesh sieve for later use.

Example 2

(1) Wash and dry the picked feijoas, peel and cut them into pieces, then put them into deionized water at 95° C. and boil them for 5 minutes to inactivate pectinase. After properly controlling the water, put it in a blast drying oven at 55°C and dry it until the water content is less than 6%. After pulverizing with a pulverizer, pass through a 60-mesh sieve to make feijoa pulp powder (pulp and pulp) and seal it in a self-sealing container. bag, and store in a desiccator for later use.

(2) Take 10g of feijoa powder from step (1) in a 250mL conical flask, add 160mL of 50mM sodium citrate buffer (solid-to-liquid ratio 1:16), adjust the pH to 6.8 with citric acid, and mix well.

(3) Put the mixture of (2) into a microwave oven (MM823ESJ-PA Midea microwave oven), set the heat to medium-low heat (350W IEC) for 10 minutes, take it out and cool it until it is not hot.

(4) Weigh β-glucosidase (250U/g, the ratio of enzyme activity to substrate) in a conical flask, mix well, and carry out enzymolysis treatment in a constant temperature shaking incubator for 90min, and the enzymolysis temperature is set at 50°C, the rotating speed of the shaker is 150rpm.

(5) After the extraction, heat in a 95° C. water bath for 5 minutes to inactivate the enzyme.

(6) Centrifuge the extracted solution at 5500 rpm for 8 minutes, collect the supernatant, wash the residue twice with distilled water, and record the volume after combining the supernatant.

(7) Add twice the volume of absolute ethanol to the supernatant, mix well and let it settle for two hours, centrifuge at 5000 rpm after the pectin is precipitated, and wash the pectin in the test tube 3 times with absolute ethanol.

(8) Dry the cleaned pectin in an electric blast drying oven at a set temperature of 50° C., grind it with a mortar and pass it through a 100-mesh sieve for later use.

Example 3

(1) The plucked feijoas were washed, dried, peeled and cut into pieces, then placed in deionized water at 90° C. and boiled for 5 minutes to inactivate pectinase. After properly controlling the water, put it in a blast drying oven at 50°C and dry it until the water content is less than 6%. After pulverizing with a pulverizer, pass through a 60-mesh sieve to make feijoa pulp powder (pulp and pulp) and seal it in a self-sealing container. bag, and store in a desiccator for later use.

(2) Take 10g of feijoa powder from step (1) in a 250mL conical flask, add 140mL of 50mM sodium citrate buffer (solid-to-liquid ratio 1:14), adjust the pH to 6.8 with citric acid, and mix well.

(3) Put the mixture of (2) into a microwave oven (MM823ESJ-PA Midea microwave oven), set the heat to medium-low heat (400W IEC) for 9 minutes, take it out and cool it until it is not hot.

(4) Weigh β-glucosidase (250U/g, the ratio of enzyme activity to substrate) in a conical flask, mix evenly, and carry out enzymolysis treatment in a constant temperature shaking incubator for 100min, and the enzymolysis temperature is set at 45°C, the rotation speed of the shaker is 150rpm.

(5) After the extraction, heat in a 90° C. water bath for 5 minutes to inactivate the enzyme.

(6) Centrifuge the extracted solution at 5000 rpm for 10 min, collect the supernatant, wash the residue twice with distilled water, and record the volume after combining the supernatant.

(7) Add twice the volume of absolute ethanol to the supernatant, mix well and let it settle for two hours, centrifuge at 5500 rpm after the pectin is precipitated, and wash the pectin in the test tube 3 times with absolute ethanol.

(8) Dry the cleaned pectin in an electric blast drying oven at a set temperature of 50° C., grind it with a mortar and pass it through a 100-mesh sieve for later use.

Comparative example 1

This comparative example provides a kind of method that biological enzymatic method extracts feijoa pectin, comprises the following steps:

Accurately weigh 10g of feijoa pulp powder into an Erlenmeyer flask, add 150mL (1:15) of 50mM sodium citrate solution with a pH of 6.8, then add β-glucosidase (250U/g) for enzymatic hydrolysis for 99min, and enzymatic hydrolysis The temperature was set at 50° C., and the rotation speed of the shaker was 150 rpm. Other treatments are the same as in Example 1.

Comparative example 2

This comparative example provides a kind of method that conventional hot acid water method extracts feijoa pectin, comprises the following steps:

(1) Weigh 10 g of feijoa pulp powder into a beaker, add 150 mL of hydrochloric acid solution with a pH of 1, and extract in a heat-collecting magnetic stirrer with a set temperature of 95 °C for 130 min.

(2) After the extraction is completed, a centrifuge is used to separate the pulp residue from the soluble compounds, and the separated pulp residue is washed with distilled water.

(3) Collect the filtrate, centrifuge at 5000 rpm for 10 min to remove fine powder, add 2 times volume of absolute ethanol to precipitate pectin for 2 hours.

(4) The precipitated pectin was centrifuged at 5000rpm for 10min, washed with absolute ethanol for 3 times and centrifuged again, and the finally obtained pectin was dried at 50°C in an electric blast drying oven.

Test Example 1-Monosaccharide Determination

The determination method of monosaccharide adopts trichloroacetic acid (TFA) method, accurately weighs 10mg of polysaccharide sample, adds 1mL TFA (4M) solution and hydrolyzes at 110 ℃ for 4 hours. Subsequently, the hydrolyzate was co-distilled several times with ethanol under reduced pressure to completely remove TFA. Hydrolyzed samples and standard monosaccharides were acetylated by adding a mixture of hydroxylamine hydrochloride, pyridine, and acetic anhydride. The Shimadzu GC/MS gas chromatography (QP 2010Plus) system was used for analysis. The initial temperature was set at 150°C and kept for 17 minutes. The temperature program increased from 150°C to 165°C at a rate of 1°C/min, then increased to 200°C at a rate of 10°C/min, and then increased at a rate of 50°C/min. Rise to 380°C. Sample injection volume 1μL, split ratio 1:20, carrier gas helium, flow rate 1mL/min; ion source temperature 230°C, electron impact voltage 70eV, mass scanning range m/z20-650, frequency 0.2s/scan, solvent The delay time is set to 5.5min. Standard quantification was performed with β-phenyl-glucoside (0.05% w/v) as the internal standard, and the RG-I content was calculated according to the following formula:

RG-I(%)=2Rha(%)+Ara(%)+Gal(%)

Test example 2-determination of degree of esterification (DE)

The determination method of the degree of esterification is carried out with reference to the titration method in the national food safety standard GB25533 2010.

Test Example 3 - Determination of Rheological Properties

Pectin samples were dissolved in deionized water to make a concentration of 1.0% (w/v). The rotational rheometer is a Haake RS6000 owned by American Thermo Company. The probe uses a flat circular steel plate (diameter 60 mm), and the gap is set at 1.0 mm to measure the apparent viscosity of the sample. The apparent viscosity curve was measured at 25°C with the shear rate gradually increasing from 0.1 s ^-1 to 100 s ^-1 .

The experimental results are shown in Table 1 and Figure 1.

Table 1 Physical and chemical parameters of pectin

It can be seen from Table 1 and Figure 1 that the extraction method disclosed in the present invention can greatly increase the yield of pectin, and maintain a relatively high level of esterification degree and RG-I content. The extraction method disclosed by the invention can retain more RG-I domain structure of pectin, and extract high-ester pectin rich in RG-I. At the same time, the degree of esterification of pectin extracted by the invention is 68.43%, which belongs to high-ester pectin, and its viscosity is equivalent to that of pectin extracted by enzymatic method, which is better than that of pectin extracted by conventional hot acid water method.

Although the specific implementation of the present invention has been described in detail, it should not be construed as limiting the protection scope of this patent. Within the scope described in the claims, various modifications and deformations that can be made by those skilled in the art without creative work still belong to the protection scope of this patent.

Claims (10)

1. an extraction method of feijoa pectin rich in RG-1 high ester, is characterized in that, comprises the following steps: (1) Boiling the feijoa pulp and drying it until the water content is lower than 6%, and mixing the powder and the buffer evenly after crushing; (2) Microwave heat treatment of the mixed mixed liquid followed by enzymatic hydrolysis, centrifugation and purification, followed by drying the pectin; The power of the microwave heating treatment is 350-400W, and the time is 5-15min. 2. the extraction method of the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1, is characterized in that, the temperature of described step (1) boiling is 85-100 ℃, and the time of boiling is 3 -8min. 3. the extraction method of the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1, is characterized in that, the temperature of described step (1) drying is 45-55 ℃. 4. the extraction method of the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1, is characterized in that, the powder after described step (1) pulverizes crosses 60 mesh sieves. 5. the extraction method of the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1 or 4 is characterized in that, the ratio relation of described step (1) powder and damping fluid is 10g: 120- 180 mL, and the buffer solution is a 50 mM sodium citrate buffer solution with a pH of 6.8. 6. the extraction method of the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1, is characterized in that, described enzymolysis process is specially: with β-glucosidase and feijoa powder according to After the amount of 250U/g is mixed evenly, the enzymolysis treatment is carried out for 90-100min, the temperature of the enzymolysis is 45-55°C, and the rotation speed of the enzymolysis is 120-180rpm. 7. the extraction method of the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1, is characterized in that, after described step (2) enzymolysis, the solution after extraction is at 90-100 ℃ Heat in a water bath environment for 3-5min. 8. the extraction method of the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1, is characterized in that, the centrifugal rotating speed of described step (2) is 4500-5500rpm, and the time of centrifugal is 8- 12min. 9. the extraction method of the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1, is characterized in that, described purification specifically comprises the following steps: collect the supernatant after centrifugation, in supernatant Add twice the volume of absolute ethanol, mix well and let it settle for 2 hours, centrifuge at 4500-5500rpm after the pectin is precipitated, and wash the pectin in the test tube 2-3 times with absolute ethanol. 10. the method for extracting the feijoa pectin that is rich in RG-1 high ester as claimed in claim 1, is characterized in that, the temperature of described pectin drying is 45-60 ℃.

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