KR100479359B1 - New microorganism degrading seaweeds - Google Patents

Abstract

The present invention has an excellent effect on the decomposition and fermentation of seaweeds such as seaweed, seaweed, seaweed, seaweed, butts, etc., and is a novel microorganism DS-01 {depository institution: Biotechnology Research Institute Genetic Bank (KCTC); Accession No. SD1006} and a degrading enzyme isolated from the microorganism. New strain DS-01 maximizes the efficiency of seaweeds by rapidly degrading and degrading various seaweeds, and thus can be used in various fields including the manufacture of health and beauty drinks and functional feeds containing active ingredients of seaweeds.

Description

New seaweed decomposition {NEW MICROORGANISM DEGRADING SEAWEEDS}

The present invention relates to a novel microorganism having an excellent effect in degrading algae and algae degrading enzyme isolated from the microorganism, and more particularly, to decompose and ferment algae such as seaweed, seaweed, seaweed, seaweed, wormwood, etc. New microorganism, DS-01 {depository: Biotechnology Research Institute Genetic Bank (KCTC); Accession No. SD1006} and a degrading enzyme isolated from the microorganism.

       Seaweeds, including seaweed, are the most abundant in alkalis, but contain few calories, and contain many minerals and nutrients essential to our body, such as vitamins A, B1, B2, and C. It is now attracting much attention as one of the ideal foods to provide nutritional balance for modern people with severe nutritional imbalances.

In particular, many pharmacological effects are known for alginic acid, which is a polysaccharide of marine plants that is comparable to cellulose of land plants and occupies about 30% of its constituents. For example, alginic acid promotes the movement of the stomach to effectively prevent constipation, easily remove stool, and stays in the large intestine or small intestine to reduce fat and cholesterol, which causes adult disease, cholesterol prevention and heavy metals, It combines with pollutants such as pesticides and has the effect of being discharged out of the body. In addition, not only inhibits the production of harmful oxygen known to promote aging, but also increases the enzyme to remove it, it is known to have an anti-cancer effect by preventing the cancer cell process of normal cells. In addition, alginic acid is a calorie-free and also known to prevent obesity, and many examples of using it as a food material have been reported.

It is known that the pharmacological effect of such algae is significantly increased when the molecular weight of algae and other algae is degraded.

As a method of reducing the molecular weight of the seaweed fiber, a method based on an aqueous acid solution and a method of applying heat are generally present. However, this not only has the problem that many minerals, the main functional ingredients of seaweeds, are leached into solution, but they are also produced in large quantities during a short production period and are processed quickly before decaying algae several times faster than land plants. In order to solve this problem, a large-scale treatment facility is required.

On the other hand, a method of low molecular weight polysaccharides by irradiation with ultraviolet rays, gamma rays, electron beams, or ultrasonic waves has been reported, but this method also has a problem as described above in treating a large amount of algae produced in a short production period. I still have it.

Accordingly, the present inventors have studied the seaweed microorganisms and the method of using the degrading enzymes to decompose and ferment algae as a solution to solve the above problems and to simultaneously solve the problem of low molecular weight and storage of algae, and as a result The present invention has been completed by identifying and identifying new strains having excellent decomposition efficiency.

Accordingly, it is an object of the present invention to provide a new strain having excellent algal resolution and algae degrading enzyme isolated therefrom.

In order to achieve the object of the present invention as described above, the present invention is a new strain of a new genus belonging to Gamma-proteobacteria DS-01 {depository institution: Biotechnology Research Institute Gene Bank (KCTC); Accession No .: SD1006}.

In addition, the present invention is characterized in that it provides an algae degrading enzyme produced from the new strain DS-01.

Hereinafter, the isolation, identification of the novel strain DS-01 according to the present invention and the properties of the substance secreted by the microorganism will be described in more detail.

1. Sampling and Strain Separation

In April 2001, samples were taken from the southern part of the South Coast. After dividing the collected 약 by about 10g, each of them was dissolved in 200ml of sterile distilled water and left to settle. The separated solution was mixed with 10 g of seaweed cut to about 2 cm into a culture solution containing 2.5 g of pepton and 0.5 g of yeast extract in 500 ml of seawater and shaken. After 48 hours, the size of the seaweed sample was reduced to about 1 to 3 mm from the first 2 cm, 0.1 ml of this culture was plated on the marine agar solid medium. Six kinds of bacteria were cultivated when incubated for 3 days in a constant temperature and humidity chamber. Among them, marine bacteria isolate and decay the solid medium, which is called DS-01.

2. Strain Identification

Identification of the purely isolated DS-01 strain was requested to MicroID Co., Ltd. As a result of the identification, the DS-01 strain was determined to be a new genus belonging to Gamma proteobacteria (FIGS. 1A to 1E).

This strain was named DS-01, and was deposited on July 9, 2001 under the accession number SD1006 to the Biotechnology Research Institute Gene Bank (KCTC).

Table 1 shows the basic characteristics of the DS-01 strain.

 Characteristics of the present invention DS-01   1. Optimum growth temperature 20 ~ 30 ℃  2.Concentration of NaCl (%) 0.5 1 3 5 10 20 Growth-+++- Catalase Activity - Hydrolytic Properties: Urea Starch Casein Gelatin ---- Carbon source utilization characteristics: sucrose cellobiose xylose xylose mannitol fructose galactose glucose lactose rhamnose maltose maltose ++-+++++++

Experimental Example 1 Alginate Degradation Capacity of DS-01

200 ml of culture solution (hereinafter referred to as basic culture solution) prepared by dissolving 5 g of peptone and 1 g of yeast extract in 1 liter of seawater, 10 g of seaweed leaf cut to about 2 cm size, and about 0.1 g (dry weight) of DS-01 bacteria 10 ml of the culture solution was mixed, digested for 60 hours, filtered, washed, and dried. Alginic acid was extracted and separated from the dried wakame leaves with 0.3 wt% Na ₂ CO ₃ aqueous solution. This was dissolved in 0.2N NaCl aqueous solution and the average molecular weight was calculated by Mark-Houwink-Sakurada equation after viscosity measurement using a viscometer (VISCOTEK CO.Model Y-500) for each concentration was 1,617. The average molecular weight of wakame before decomposition was 31,977, which showed a decrease in molecular weight of about 1/20 by decomposition for 60 hours.

Experimental Example 2 Investigation of Degradation Capacity of Seaweed Leaves of DS-01

10 g of seaweed leaves were cut to a size of about 2 cm and digested with 0.1 g of DS-01 in 200 ml of the primary culture medium at room temperature.

Size after disintegration of wakame leaves by DS-01       Size Sample 30mesh pass through% 50mesh pass% 60mesh pass% 80mesh pass% 100mesh pass through% 120mesh pass through% 140mesh pass% 170mesh pass% 200mesh Pass% 2-day decomposition 85.6 68.9 55.7 45.4 32.0 23.7 18.6 13.4 7.2 4 days decomposition 93.5 79.0 73.0 61.0 46.0 38.5 30.0 24.0 18.0

Experimental Example 3 Investigation of Seaweed Degradation Capacity of DS-01

      10 g each of kelp, seaweed, laver, sea urchin, and hearing cut into about 2 cm lengths are put into 200 ml of the basic culture solution containing about 0.1 g of DS-01 bacteria, and the samples are decomposed into small particles having a diameter of 1 mm or less. As a result of measuring the time to become as shown in Table 3.

 Disassembly of each seaweed by DS-01 bacteria (time to disassemble to less than 1mm) Kelp 톳 Kim Woodfish ear Decomposition time (days) 4 4 4 3 3

Experimental Example 4 Growth Curve of DS-01

      As a result of measuring the weight change of the bacteria while growing DS-01 bacteria in the basic culture medium at room temperature was as shown in FIG. At about 5 days, the concentration of the fungus peaked and then decreased.

Experimental Example 5 Seaweed Degrading Enzyme Induction Test from DS-01

     To 200ml of the basic culture solution containing 0.1g of DS-01 bacteria, 2.5g each of agar and alginate were put into enzyme-induced experiments. After incubation for 2 days, the bacteria were removed and 5 g of 2% aga bead was added to the remaining solution, and the weight change of the beads was measured.

 DS-01 Enzyme Induction Comparison        derivative agar alginate no unit / ℓ 185 225 155

   Herein, the enzyme activity unit was set to 1 μg / min = 1 unit of agar loss. Enzyme induction is best demonstrated in the presence of alginate substrates.

Experimental Example 6 Investigation of Seaweed Degradability of Seaweed Degrading Enzyme Induced from DS-01

5.0 g of alginate was added to 500 ml of the basic culture solution, and after 5 days of incubation, the bacteria were removed, and 7.2 g of Na ₂ SO ₄ was added to the remaining solution. The separated crystals (estimated by enzymes) were added to seawater and decomposed 3% alginate beads at room temperature, showing an enzyme activity of about 490 units / l. Herein, one unit decomposes pure alginate at 1 µg / min.

Experimental Example 7 Investigation of the Activity of Algae Degrading Enzyme Induced by DS-01

      After 3 days propagation culture in the main culture solution, the bacteria were removed by centrifugation (10,000rpm), and alginate beads were decomposed in the remaining solution for 12 hours at each temperature. It was like It is shown that the enzyme activity increases in proportion to the temperature when the temperature rises to 40 ℃.

 Comparison of activity according to temperature of DS-01 enzyme      Temperature (℃) activity 18 25 28 36 40 unit / ℓ 350 511 561 611 683

Experimental Example 8

1.6 g of Na ₃ PO ₄ · 12H ₂ O was added to 200 ml of the basic culture solution, and about 0.1 g of DS-01 was added thereto, followed by incubation at room temperature for 5 days. After adding the beads and reacting at room temperature for 12 hours, the enzyme activity was found to be 834 unit / l.

Experimental Example 9

      After mixing 0.5 g of DS-01 bacteria in 20 g of seaweed with 80% water content, the mixture was sealed and placed in an atmosphere of 30 ° C. and a relative humidity of 80%. After 3 months, there was no decay by various bacteria and only degradation of seaweed by DS-01.

Experimental Example 10

      Minerals were analyzed by atomic absorption spectrophotometer (HITACHI, Z-2000 Model) before and after the disintegration of seaweed in order to determine whether minerals in the seaweed were lost to the culture medium during the disintegration of seaweed by DS-01.

      After 5 days of disintegration of seaweed leaves by DS-01 bacteria, the concentration of Fe and Zn in seawater culture was 0 ppm, indicating that Fe and Zn in seaweed were not lost.

      Degradation of seaweed leaves for 4 days after analysis of Se (an important ingredient to prevent postpartum stagnation in cows) of seaweed leaves showed 4.1 mg in 100 g of seaweed. It was shown that there was a decrease of Se in some seaweeds before the decomposition, but the amount of loss was so small that the mineral loss was negligible in the process of seaweed decomposition by DS-01.

The new strain DS-01 of the present invention maximizes the efficiency of use of seaweeds by decomposing various seaweeds such as seaweed, kelp, seaweed, shellfish, and beetroot at a low rate, thereby lowering the molecular weight, and thus the health and beauty drinks containing the active ingredients of seaweeds and It is available in a variety of fields, including the manufacture of functional feeds.

       Figure 1a is the front page of the identification result report of strain DS-01 according to the present invention.

       Figure 1b is the second page of the identification result report of strain DS-01 according to the present invention.

       Figure 1c is a third page of the identification result report of strain DS-01 according to the present invention.

       Figure 1d is the fourth page of the identification result report of strain DS-01 according to the present invention.

       Figure 1e is a fifth page of the identification result report of strain DS-01 according to the present invention.

       Figure 2 is a graph showing the weight change according to the days of culture of strain DS-01 according to the present invention.

Claims (2)

New strain DS-01 (Accession No .: KCTC-SD1006) belonging to gamma proteobacteria with seaweed resolution. delete