KR20110089905A - An antimicrobial use of onion peels subcritical water extracts - Google Patents

Abstract

PURPOSE: A method for preparing subcritical water extract of onion skin using purified water is provided to obtain an antibacterial agent. CONSTITUTION: A method for preparing subcritical water extract of onion skin comprises: a step of mixing washed onion skin and diatomaceous earth in a ratio of 1:2-1:3 to prepare a sample; a step of adding the sample to 34ml or 66ml of a cells; a step of adding distilled water; and a step of extracting at 60 pressure and 100 degrees Celsius for 15 minutes. An antibacterial agent contains the subcritical water extract of onion skin.

Description

An antimicrobial use of onion peels subcritical water extracts}

The present invention relates to the use of the subcritical water extract of onion peel can be used as an antimicrobial agent of the subcritical water extract of onion.

Onion ( Allium cepa L.) is a perennial plant belonging to the family Liliaceae, and has a long history of cultivation and has a unique taste and aroma.

Recent studies have found that onions contain polyphenols that exhibit antimicrobial activity and sulfur compounds with various physiological activities, increasing interest as a functional food for preventing adult diseases. However, 10-30% of onions used in the food processing field of domestic agricultural products are peels, and by-products are generated annually, which are often discarded. These discarded agricultural by-products can contribute to the rural economy by increasing farm household income when used as functional materials. In particular, if the antibacterial or antioxidant functional substances can be extracted from onion peels, which are onion by-products, the rural economy can be activated.

Soxhlet extraction and solvent extraction are used for the extraction of agricultural foods and by-products. However, these extraction methods have a long extraction time and can cause secondary contamination by a large amount of solvents such as acetone, dichloromethane and hexane.

Subcritical water extraction (SWE) is a method in which a specific organic material can be extracted in a short time by changing the dielectric constant of water with temperature and pressure in a solid or semi-solid sample. In addition, the use of water as the extraction solvent can greatly reduce the organic solvent used in the separation process. Subcritical conditions refer to temperature and pressure ranges below the critical point (T = 374.2 ° C, P = 218.3 atm) where the liquid and gaseous states of water coexist. Water loses its intrinsic polarity at the pressure of 50-150 atm near the critical point and at the temperature of 200-300 ° C, and loses its intrinsic polarity, and the dielectric constant is drastically reduced. In this case, the dielectric constant of water may be 2.5 or less, and may be used as an extraction solvent for a specific organic material.

Staphylococcus aureus is widely distributed in air, soil, etc., and is particularly likely to be contaminated with foods rich in protein and carbohydrates. have. In recent years, the incidence of food poisoning caused by S. aureus has increased in countries around the world. In Korea, according to the 2003 domestic food poisoning statistics, food poisoning caused by S. aureus was 9.62%, the third after enteritis vibrio and Salmonella spp. It is known as a causative agent of causing many food poisonings.

Since hospital infections with S. aureus and methicllin-resistant Staphylococcusaureus (MRSA) have been reported in Japan in the early 1990s, 'in-hospital infection with MRSA' has emerged as a serious problem in the medical industry.

MRSA, a strain of S. aureus , was first reported in the UK in 1961 and is prevalent throughout the world and is mutated to produce β-lactamase enzymes that destroy the β-lactam structure of the antibiotic methicllin. Staphylococcus. In addition, since it is resistant to most antibiotics, it is a pathogenic infectious bacterium that currently requires countermeasures worldwide. When infected, the antibiotics currently used are difficult to treat, causing frequent deaths of patients. Therefore, there is an urgent need for the development of new antibiotics for the prevention and treatment of infections against this strain.

Republic of Korea Patent No. 10-0620568 "Extraction, Separation and Purification of Isoflavones from Soybeans Using Supercritical / Subcritical Water" Republic of Korea Patent No. 10-0626221 "Manufacturing device and method of manufacturing healthy rice using subcritical / supercritical fluid and ultrasonic wave" Republic of Korea Patent No. 10-0452467 "Method for extracting functional substance and removing oxidizing substance from citron using supercritical or subcritical fluid" Korean Patent Application No. 10-2002-0069524 "Method for Extracting Antimicrobial Material from Wasabi using Supercritical and Subcritical Fluid Extraction Processes"

Griffiths G, et. al., onion-a global benefit to health. Phytother. Res. 16: 603-615 (2002) Jakubowski H, On the health benefit of Allium sp. Nutrition 19: 167-168 (2003) Turner C, et. al., Subcritical water extraction and β-glucosidase-catalyzed hydrolysis of quercetin glycosides in onion waste. Green Chem. 8: 949-959 (2006) Ramos L, et. al., Current use of pressurized liquid extraction and subcritical water extraction in environmental analysis. J. Chromatogr. A. 975: 3-29 (2002)

Accordingly, an object of the present invention is to obtain an onion peel subcritical water extract using subcritical water extraction method (SWE) from onion peel which is a by-product generated in the food industry, to provide a use as an antimicrobial agent as an active ingredient.

The object of the present invention is the step of subcritical water extraction onion shell; It was achieved through the step of verifying the antibacterial functionality of the subcritical water extract derived from onion peel obtained from the above and evaluating its use as an antimicrobial agent.

The present invention has an excellent effect of providing an antimicrobial onion peel subcritical water extract by the problem solving means.

1 is a diagram showing a process of the preferred subcritical water extraction method of the preferred onion peel according to the present invention.
2 is a graph showing growth inhibition (antibacterial) effect on S. aureus strains of Quercetin compounds as a control experiment result according to the present invention.
Figure 3 is a graph showing the growth inhibition (antibacterial) effect on S. aureus strains of subcritical water extract of the onion peel of the present invention.

Preparation of Experimental Materials and Onion Samples

Onions used in this experiment were purchased from the market as Uiseongsan onion in Gyeongsangbuk-do. Dry the hot air at 60 ℃ using only the orange skin of onion, and then crush it into fine powder (1-10 mm) using a high speed mixer (Blender 7012S, Waring, Torrington, USA) to keep it refrigerated (4-5 ℃). It was used for the experiment.

Microbial strain

Staphylococcus aureus KCCM 40501, KCCM 3395 and KCCM 11335 were purchased from Korea Microbiological Conservation Center (Seoul). S. aureus KCCM 40510 is methicillin resistant Staphylococcus aureus (MRSA). The strain was used in the experiment after pre-culture at tryptic soy broth (Difco) to which 0.6% yeast extract (TSB-YE) was added three times for 12 hours at 37 ℃.

Ethanol extraction

20 g of onion peel and 200 mL of 70% (v / v) ethanol were mixed and extracted at 60 ° C. for 3 hours. After extraction Whatman No. It was filtered with 2 filter paper, and the filtered residue was extracted twice more under the same conditions. The extracted ethanol extract was concentrated by removing the solvent using a vacuum condenser (EYELA N-1000V, Tokyo, Japan). After concentration under reduced pressure, the extract was freeze-dried and stored frozen and used in the control experiment of the present invention.

Hydrothermal extraction

20 g of onion peel and 200 mL of 70% (v / v) distilled water were mixed and extracted at 80 ° C. for 3 hours. After extraction Whatman No. It was filtered with 2 filter paper, and the filtered residue was extracted twice more under the same conditions. The extracted hot water extract was concentrated using a reduced pressure concentrator (EYELA N-1000V, Tokyo, Japan). After concentration under reduced pressure, the extract was freeze-dried and stored frozen and used in the control experiment of the present invention.

Preparation of Subcritical Water Extract

A subcritical extractor system (DIONEX ASE 100, Dionex Corporation, Sunnyvale, California, USA) was used for onion peel extraction. Finely chopped onion skin and diatomaceous earth (DE) were properly mixed and placed in a 34 mL or 66 mL size cell, mounted in an extractor, and extracted. The preferred ratio of onion peel and DE is 1: 3 or 1: 2. The extraction solvent was extracted by setting each extraction temperature and extraction time using only Milli-Q water (Fig. 1). Extraction conditions for the antimicrobial experiment of Staphylococcus aureus were most preferably used for 15 minutes at 60 atm and 110 ℃ and quercetin as a comparative group. The water extract sample of the present invention was used while stored in a 4 ℃ refrigerator.

Antibacterial effect measurement

In order to examine the effect of subcritical onion peel extract on S. aureus growth, the inhibitory effect of growth on broth was measured. The initial bacterial count of S. aureus was adjusted to 2.6 × 10 ⁶ CFU / mL. 100μL of bacterial solution and SWE extract were inoculated in tryptic soy broth containing 0.6% yeast extract and incubated at 160 rpm and 37 ℃.

SWE onion extract was used by diluting twice, and the final concentration was adjusted to 40 ppm, 80 ppm, 160 ppm. The control group was treated with the same amount of distilled water.

The sample was sampled every 2 hours while incubating for 0-8 hours and the absorbance was measured at 630 nm using a Spectrophotometer (Optizen 2120 UV, Daejeon, Korea). Sampling samples were diluted in 0.1% peptone water and smeared on tryptic soy agar containing 0.6% yeast extract to measure the number of viable cells. The viable cell count was calculated by incubation for about 20 hours after smearing.

Quercetin was used as an antimicrobial standard, and the final concentration was adjusted to 25, 50, 100, 200 μM and the same experiments as above (Fig. 2).

Polyphenols containing flavonoids are known to have antimicrobial effects. The mechanism of these active ingredients is not known exactly, but most are known for their inhibition of nucleic acid synthesis, plasma membrane function, and energy metabolism.

In the present invention, quercetin was used as a positive control to measure the growth inhibitory effect of SWE extract against various strains of S. aureus (FIG. 3). This is because the major flavonoids present in large quantities in onion peels are known as quercetin. As a result of Figures 2 and 3, the SWE extract was inferior in antibacterial effect than pure quercetin. Quercetin 200 μM treatment group showed low bacterial growth rate of 1.0-1.5 log cycle after 8 hours. On the other hand, SWE extracts showed 1.6-1.9 log cycles, which were higher than quercetin after 8 hours. However, it showed lower bacterial growth rate than the control without SWE extract. As a result, it was observed that the SWE onion extract had about 2.3 times lower antibacterial activity than quercetin when converted to the same concentration. It is believed that SWE extract is not 100% quercetin and in addition, glycosides are used as nutrients in the growth of S. aureus strains.

Total polyphenol content determination

Phenol content was colorimetrically determined according to the Folin-Denis method. 0.1 mL of constant dilution extract and 2 mL of 2% Na ₂ CO ₃ were mixed, left at room temperature for 3 minutes, and then 0.1 mL of Folin-Ciocalteau reagent (Sigma Chemical Co., St. Louis, MO, USA) was added. . After mixing 30 minutes, the absorbance was measured at 750 nm using a spectrophotometer. The total phenol content of the sample extract was calculated from the standard calibration curve obtained by preparing quercetin as a standard.

The total polyphenolic compound is analyzed by measuring the reducing power of the extract against the Folin reagent. As a result of Table 1, the total polyphenol content of the onion peel extract used in the present invention showed a significant difference depending on the extraction method. The total polyphenol content of onion skins is 26.68-372.5 mg quercetin / g extract.

The total polyphenol content per gram of extract was highest in ethanol extract (372.5 mg quercetin / g extract). However, the content of 1 g of onion peel raw material was highest in the subcritical extract extracted at 110 ° C. for 15 minutes (38.84 mg quercetin / g onion peel).

The content of SWE extraction was higher in the 110 ℃ subcritical extract than in the 165 ℃ subcritical extract. It is believed that the active ingredient of onion peel is destroyed and decomposed because 165 ℃ subcritical extract is extracted at high temperature.

As described above, the subcritical water extract performed at 60 atm and 110 ° C. for 15 minutes is an extremely useful invention in the biopharmaceutical and food processing industries because it has an excellent effect of providing use as an antimicrobial agent.

Claims (1)

Prepare the sample by combining 1: 2 or 1: 3 of finely divided onion skin and diatomaceous earth, put it in a 34 mL or 66 mL size cell, and add distilled water for 15 minutes at 60 atm and 110 ° C. Antimicrobial agent with subcritical water extract of onion peel as an active ingredient.

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