KR101631057B1 - Method for extracting carotenoid from fermented persimmon sludge - Google Patents

Abstract

The present invention relates to a method for extracting carotenoids from a selected one of persimmon sludge fermented with microorganisms, extract of persimmon sludge fermented with microorganisms or cultures obtained by culturing microorganisms in sensory sludge, It is possible to reduce the disposal cost of the sludge disposal, and it is possible to extract carotenoids directly from the bacteria cultivated in the sludge together with additional profit generation, because the sludge containing the sludge can be recycled and utilized as high value-added biomaterial The production cost can be reduced.

Description

Technical Field [0001] The present invention relates to a fermented persimmon sludge,

The present invention relates to a method for producing carotenoids from fermented persimmon sludge.

It is rich in sugars such as glucose and fructose, and acts as an energy source for lactic acid bacteria. It is rich in vitamin A, vitamin C, minerals or dietary fiber, and has a unique natural coloring and aroma component. It is also known that persimmon has a variety of physiological activities such as antioxidant, anti-arteriosclerosis, anticancer, stimulation of alcohol metabolism, anti-blood coagulation and antiallergic.

As of 2011, Korea has the second largest production capacity with 390,820 M / T in domestic production, and its production is steadily increasing to 219,124 M / T.

It is processed and consumed in various forms such as frozen season, half-dry, three-half-dry, wine-making and mulberry-dyeing. It has excellent practicality such as ultraviolet light, light blocking, good ventilation and increased durability after dyeing without any additional processing. Therefore, in Qingdao, only about 30 companies have been added to the dyeing industry, and 600 M / T per year is processed for dyeing. However, due to the development of this dyeing business, a large amount of juice sludge having a concentration of about 240 M / T is generated, which is partially used as feed, fertilizer, or as industrial waste. As the nutrients and effective ingredients remain in the juice sludge, they are lost when they are discarded as they are, and because they are high in water and organic matter, they are easily decomposed and cause environmental pollution. ) Costs of 30-50 million won per day.

In order to solve the above problems, there is a demand for research on extracting functional materials from discarded sludge and reusing them or commercializing them using extracts.

In Korean Patent No. 10-0853313 (Patent Document 1), there is disclosed a method for extracting beta-carotene from a sensory and fatigue which is a by-product produced in a persimmon processing step. However, the method of extracting beta-carotene from persimmon flesh, There has been no report on a method for producing a functional substance in a contained sludge.

1. Domestic registered patent 10-0853313

It is therefore an object of the present invention to provide a process for producing carotenoids from garlic sludge which is discarded after squeezing.

In order to achieve the above object, the present invention provides a method for producing carotenoids, which comprises extracting carotenoids from a selected one of a sensed sludge fermented with a microorganism, an extract of a sensed sludge fermented with a microorganism or a cultured product obtained by culturing a microorganism in a sensory sludge ≪ / RTI >

The present invention provides a method for extracting carotenoids from a selected one of a sensed sludge fermented with a microorganism, an extract of a sensed sludge fermented with a microorganism or a cultured product obtained by culturing a microorganism in a sensory sludge, Can be recycled and utilized as high-value-added biomaterials.

FIG. 1 is a graph showing the content of total phenol in sensory sludge extracted with water or ethanol.
2 is a graph showing the content of total carotenoids in sensory sludge extracted with water or ethanol.
FIG. 3 is a graph showing the content of reducing sugar in the reducing sludge extracted with water or ethanol.
4 is a graph showing the content of total carotenoids in the sludge extracted with water (DW), ethanol (E), methanol (M), acetone (A), hexane (H), chloroform (C) and DMSO .
FIG. 5 is a graph showing the total carotenoid content of the extract of the sludge according to the degree of grinding. The sludge was extracted with water (A), ethanol (B) and methanol (C) , Ethanol (E) and methanol (F). The dried sludge was extracted with water (G), ethanol (H) and methanol (I).
Fig. 6 is an image showing sensed sludge fermented with Rhodotorula aurantiaca, Rhodotorula glutinis, Rhodotorula rubra, and Rhodotorula radicillinosa.
FIG. 7 is an image showing the contents of carotenoids obtained by lyophilization and pulverization of Rhodotorula glutinis, Rhodotorula lubura, and Rhodotorula mushroom Rhizoma cultured in a sludge sludge.
8 is an image showing carotenoids obtained by lyophilization and pulverization of Rhodotorulaglutinus cultured in a sludge sludge.

The inventors of the present invention have found that, in order to utilize the sensed sludge produced after the persimmon extract, the carotenoid is extracted from a selected one from a sensate sludge fermented with a microorganism, an extract of a sensate sludge fermented with a microorganism or a culture obtained by culturing a microorganism in a sensate sludge And the optimum extraction conditions for the extraction were investigated to complete the present invention.

Carotenoids are lipophilic pigments with orange, yellow or red color, and are known to act as a powerful remover for light energy absorption, oxygen transfer and singlet oxygen as precursors of vitamin A (vitamin A).

The sensory sludge may be selected from the group consisting of perspiration sludge, sausage sausage sausage, and dried sausage sludge. Preferably, the sausage sausage is dried and crushed, but the present invention is not limited thereto.

The drying may be carried out by a natural circulation drying method or a freeze drying method, preferably by a natural circulation drying method, but is not limited thereto.

In addition, the present invention provides a method for producing a sensory sludge extract, comprising the steps of: inoculating 100 parts by weight of a total amount of sensory sludge and inoculating the microorganism with 1 to 10 parts by weight; And extracting the fermented fermented product with a solvent.

The microorganism may be any one selected from the group consisting of Rhodotorula aurantiaca, Rhodotorula glutinis, Rhodotorula rhubarb, and Rhodotorula rhodigrinosa, but the present invention is not limited thereto.

Preferably, the fermentation is carried out at 25 캜 to 35 캜 for 24 to 48 hours.

The extraction solvent may be any one selected from the group consisting of acetone, C1-C4 alcohol, dimethylsulfoxide (DMSO), water, and mixtures thereof, but acetone may be preferably used.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, preferred embodiments of the present invention will be described in detail with reference to the accompanying drawings. However, the following examples illustrate the contents of the present invention and are not intended to limit the scope of the invention.

< Reference Example > Sense Sludge  Ready

The sensory sludge, which is the residue produced, was supplied and received frozen (Qingdao County) and used for the experiment.

< Example  1> Sense Sludge  Preparation of extract

The sludge was extracted with water or ethanol as follows. When extracted with water, 5 parts by weight of distilled water was added to the sludge, and the mixture was extracted on a water bath at 65 ° C for 24 hours, filtered and lyophilized. When extracted with ethanol, 5 parts by weight of ethanol was added to the sludge, and the mixture was repeatedly cooled by refluxing for 12 hours in a water bath at 65 ° C for 2 hours. After that, the extract was filtered, reduced in pressure and concentrated, lyophilized, It was used in this experiment.

&Lt; Example 2 & gt; Screening of active ingredients of persimmon sludge

1) Sense Sludge  Determination of total phenol content

The content of phenolic compounds in the sludge was measured using Folin-Ciocalteu reagent (FCR). At this time, the Folin-Ciocalteu reagent (FCR) produces phosphotungstomolydbdic acid as a mixed solution of phosphomolybdate and phosphotungstate, When phosphotungstomolydbdic acid reacts with a reducing agent in an alkaline state, it is reduced to a color change in blue, and the amount of polyphenol can be quantified by measuring the absorbance thereof.

The total phenolics content of the total phenolic compounds was measured by a known method (J. Biol. Chem. 12: 239-243 (1912); Nutr. 35: 1309-1314 (2006) The lyophilized sample was dissolved in distilled water to adjust its concentration. 1 mL of Folin-Ciocalteu reagent (Sigma, USA) was added to 0.2 mL of the sample, and the mixture was reacted at room temperature for 3 minutes. 0.8 mL of 10% Na ₂ CO ₃ was added to the reaction solution, reacted at room temperature for 1 hour, and absorbance was measured at 765 nm. At this time, the total phenolic compound content was calculated from a standard curve (Y = 0.0162 x 0.0917, R ² = 0.9998) using gallic acid (Sigma, USA).

As a result, as shown in Fig. 1, the water extract and ethanol extract of persimmon sludge had polyphenol contents of 32.59 μg / mL and 70.33 μg / mL, respectively. Therefore, it was confirmed that more phenol could be extracted by using ethanol than water in sensory sludge extraction.

2) Sense Sludge  Determination of total carotenoid content

The total carotenoid content of the sludge was analyzed by measuring the absorbance at 450 nm using the spectrophotometric method of the already known method (Molecular & General Genetics. 255: 19-27 (1997)). At this time, the total carotenoid content was calculated from a standard curve (Y = 0.0981 x 0.0503, R ² = 0.9999) using the standard product? -Carotene dissolved in chloroform.

As a result, as shown in Fig. 2, the water extract and the ethanol extract of the sludge had carotenoid contents of 0.18 μg / mL and 6.79 μg / mL, respectively. The carotenoid content of the ethanol extracts was much higher than that of the water extracts. It was judged that carotenoids were extracted more from ethanol than water because they were lipid soluble substances.

3) Sense Sludge  Determination of the content of reducing sugar

Reducing sugar was determined by 3-amino-5-nitrosalicylic acid (3,5-dinitrosalicylic acid, DNS) reduced by reducing sugar according to the reducing sugar method (DNS method) -nitrosalicylic acid was measured. 0.5 mL of the sample solution was added with 0.5 mL of the DNS solution, followed by reaction in boiling water at 100 ° C. for 5 minutes, followed by cooling in cold water. Then, 2 mL of distilled water was added to the reaction solution, and the absorbance was measured at 546 nm. At this time, the content of reducing sugar was calculated from a standard curve using glucose (Y = 0.0014 x 0.0625, R ² = 0.9979).

As a result, as shown in FIG. 3, the water extract and the ethanol extract of the persimmon sludge had a reducing sugar content of 722.74 μg / mL and 715.12 μg / mL, respectively, and it was confirmed that a considerable amount of reducing sugar was contained in the reducing sludge. Therefore, it was confirmed that the sensory sludge can be effectively utilized for microbial fermentation.

< Example  3> Persistence through fermentation using microorganisms Sludge  Increase, separate and purify carotenoid contents

1) Sense Sludge  Determination of Carotenoid Content in Solvent Extracts.

To investigate the carotenoid content of the extract according to the solvent, 10 mL of distilled water, ethanol, methanol, acetone, hexane, chloroform and DMSO were added to 2 g of the sludge, and the mixture was centrifuged at room temperature. Then, the absorbance of the supernatant was analyzed according to the method 2) of Example 1 above.

As a result, carotenoid contents of acetone extracts were the highest at 1.839 μg / mL as shown in FIG. 0.955 μg / mL of ethanol extract, 0.520 μg / mL of methanol extract, 0.479 μg / mL of DMSO extract and 0.197 μg / mL of distilled water extract.

2) Sense Sludge Degree of polish  Extract Carotenoid content measurement

First, the sludge was washed and the water was removed. The dried sludge was dried using a natural circulation dryer (T-700, Taein Company) at 40 ° C for 18 hours and dried using a pulverizer. Then, the sludge was lyophilized by vacuum freeze drying method using a vacuum freeze dryer (free zone 2.5L freeze dry system, labcom co.) To pulverize the sludge. Compared with the two drying methods, the natural circulation drying time is shorter than that of the freeze drying method, and the natural circulation drying method is considered to be better because the cost is low.

To investigate the content of carotenoids according to the natural circulation type dryness, 30 g of distilled water, ethanol and acetone were added to 3 g of the sludge, and the mixture was centrifuged at room temperature and the supernatant was taken. Then, the absorbance of the supernatant was analyzed according to the method 2) of Example 1 above.

As a result, the dried carrot sludge was crushed and extracted with ethanol (4.16 ± 0.24 μg / mL) or methanol (4.13 ± 0.52 μg / mL), showing the highest carotenoid content. It is thought that it is possible to extract more carotenoids by enlarging the surface area by grinding or drying and grinding after extracting rather than extracting the sludge as it is.

3) Persimmon fermented by microorganism Sludge  Carotenoid content measurement

In order to ferment the sludge as a microorganism, 100 g of the sludge was ground with 300 mL of distilled water, and then 3% of the total volume of the sludge was inoculated and cultured at 25 DEG C and 150 rpm for 48 hours. The strains of Table 1 below were used for the experiment and were distributed at the Korean Microorganism Conservation Center.

KCCM  Strain 50681 Rhodotorula aurantiaca (Rhodotorula aurantiaca) 60525 Rhodotorula glutinis (Rhodotorula glutinis) 50121 Rhodotorula rubra (Rhodotorula rubra) 50124 Rhodotorula mucilaginosa Rhodotorula mucilaginosa

As a result, as shown in FIG. 6, in the case of persimmon sludge inoculated with Rhodotorula glutinis, Rhodotorula rubra, and Rhodotorula radicillinosa, the red coloring bacteria showed remarkable color change compared to the control group there was. Also, the sludge sludge was significantly reduced and the viscosity of the fermented material increased. In the case of Rhodotorula aurantiaca, the color of the persimmon sludge fermented product was soft because the bacterial color was soft and slow in growth rate.

4) Sense Sludge In a culture medium  Determination of carotenoid content in cultured microbial cultures

Then, in order to extract carotenoids from Rhodotorula glutinis, Rhodotorula rubra, or Rhodotorula mushiensis cultures cultured using the sperm sludge as a culture medium, the fungi were lyophilized and pulverized with a mortar, Extracted with acetone and centrifuged to take the supernatant.

As a result, it was confirmed that carotenoids were also extracted from Rhodotorula glutinis, Rhodotorula rubra, or Rhodotorula mushroom Rhizoma cultures cultured using the sperm sludge as a culture medium as shown in Fig.

In order to investigate the efficiency of the extraction solvent, roto-toxin cultures were lyophilized, and then saccharified with a barbara and ethanol or acetone was added thereto to extract carotenoids.

As a result, a large amount of carotenoid was extracted when extracted with acetone rather than ethanol in Rhodotorulaglutinus cultures cultured using a sensory sludge as a culture medium as shown in FIG.

In summary, in order to extract carotenoids from the sensory sludge, carotenoid extraction by drying the sensuous sludge was higher than that of the non-dry sensory sludge, and extraction with ethanol or acetone was more efficient than using other solvents Carotenoids were extracted.

And Rhodotorula sp . ), It was considered to be the best method to extract fermentation sludge by drying and crushing with ethanol or acetone.

In addition, rhodotorula ( Rhodotorula ) cultured using a sensory sludge as a culture medium sp . ) If it is desired to extract the greatest number of carotenoids from the cultures, it is considered that the best method is to extract the carotenoids by freeze-drying the cultured strains, and then extracting them with ethanol or acetone.

Claims (8)

delete delete Inoculating 1 to 10 parts by weight of the microorganism with respect to 100 parts by weight of the dried sludge; And
Extracting the fermented fermented product with ethanol or acetone,
Wherein the microorganism is selected from the group consisting of Rhodotorula aurantiaca, Rhodotorula glutinicus, Rhodotorulla lubra, Rhodotorula lucilus, Rhodotorula lucilis, Lt; RTI ID = 0.0 &gt; carotenoid &lt; / RTI &gt; delete The method of claim 3,
Wherein the drying is carried out by a natural circulation drying method or a freeze drying method. delete The method of claim 3,
Wherein the fermentation is carried out at 25 캜 to 35 캜 for 24 to 48 hours.

delete

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