KR102181470B1 - Powder of the black soldier fly larva bred using dropwort feed containing germanium and composition for anti-oxidant and anti-cancer comprising the same - Google Patents

Abstract

The present invention provides a powder of larvae, extracts thereof, and a pharmaceutical composition, food composition, and cosmetic composition for the treatment of antioxidants and cancers comprising the same, and extracts of larvae in Dongae bred using a feed containing parsley cultivated hydroponically with germanium water. In addition, the present invention provides an antioxidant fertilizer additive and feed additive including larva powder and fecal soil in Dongae, etc. containing germanium.
The powder of larvae and the antioxidant and anticancer composition comprising the same, and the powder of larvae raised using the parsley feed containing germanium of the present invention exhibits excellent antioxidant activity and cancer cell growth inhibitory effect, so that it is a natural antioxidant or natural anticancer agent in the field of medicine and pharmacy. It can be used as, and can be usefully used as an antioxidant in the agricultural industry and cosmetics field.

Description

Powder of the black soldier fly larva bred using dropwort feed containing germanium and composition for anti-oxidant and anti-cancer comprising the powder of larvae in Dongae, etc. bred using a parsley feed containing germanium the same}

The present invention relates to a powder of larvae and an antioxidant and anticancer composition comprising the same, and more particularly, to a composition for hydroponic cultivation with germanium water, and bred by mixing the water parsley in the feed with germanium water. It relates to a powder of larvae in Dongae et al. and a pharmaceutical composition for antioxidant and anticancer, functional food composition, and cosmetic composition comprising the same.

Many insects, including Hermetia illucens , have an innate immune mechanism that expresses strong antibacterial peptides against the invasion of pathogens through long evolution even in environments that are unfavorable for survival, so it is a natural antibiotic that can replace chemical antibiotics. It is known as an excellent material.

Dongae and other larvae have been attracting attention as environmental cleansing insects with excellent decomposition ability for food wastes of humans and livestock. In this organic waste, a number of larvae are fed together, generating high temperatures (50 ~ 60 ℃) and secreting antibiotics. As a result, it has shown the industrial applicability in terms of production of pharmaceuticals and pesticide substances, such as inhibiting the access of other living organisms.

Germanium (Ge) can be classified into inorganic germanium bound to oxygen, chlorine, ammonia, etc. in nature and organic germanium bound to carbon. Organic germanium includes biosynthetic organic germanium fermented by yeast with inorganic germanium, and more than 100 natural organic germanium, including wild ginseng, ginseng, garlic, etc., that plants such as wild ginseng, ginseng, and garlic absorb inorganic germanium in the soil and contain it in the living body There are chemically synthesized organic germanium species. While long-term use of synthesized inorganic germanium (GeO ₂ ) is known to cause anemia, impaired kidney function, muscle disorders and neuropathy, organic germanium has anti-tumor effect, anti-mutagenic effect, immunity strengthening effect, virus It is known to have various pharmacological effects such as the therapeutic effect of infection, anti-inflammatory effect, antipyretic analgesic effect, excretion of heavy metals and increased motility.

Recently, some researchers have developed a cultivation method for functional crops such as parsley and rice containing organic germanium grown with inorganic germanium fertilizer. However, the development of a differentiated method that can produce high content organic germanium from plants in which inorganic germanium is absorbed is required as a method for effectively converting inorganic germanium into organic germanium by absorbing inorganic germanium into plants has not yet been developed. do.

Korean Patent Application Publication No. 10-2015-0048424 (2015.05.07.)

Kwanho Park et al. J of Seric Entomol Sci 2014, 52: 142-146 Myeong-Seon Lee. JKEIA, 2016 Dec, 10(6): 451-459

The inventors of the present invention were studying a method for producing high content of organic germanium using insects such as Dongae et al., while the germanium content in larvae and fecal soils bred using parsley containing germanium as a compound feed. It was found to have increased significantly. In addition, the extract of larvae containing such a high content of germanium and the extract of fecal soil as a product thereof exhibited antioxidant and anticancer efficacy.

Accordingly, an object of the present invention is to provide a powder of larvae in Dongae, etc. bred using a parsley feed containing germanium, and an antioxidant and anticancer composition comprising the same.

According to an aspect of the present invention, a powder of larvae (black soldier fly larva powder) is provided on Dongae, etc. bred using a feed including water parsley grown hydroponically with germanium water.

In one embodiment, the water parsley may be included in the feed as parsley: wet feed = 0.1 to 20: 100 (g/ml).

In one embodiment, the larvae may be bred for 5 to 10 days, such as Dongae.

According to another aspect of the present invention, an extract of powdered larvae is provided on the larvae obtained by extracting the powder of larvae on the larvae.

In one embodiment, the extract may be one or more selected from the group consisting of ethanol extract, methanol extract, acetic acid extract, and mixtures thereof.

According to another aspect of the present invention, there is provided a pharmaceutical composition for antioxidant or cancer treatment comprising an extract of larva powder in the Dongae and the like.

In one embodiment, the pharmaceutical composition may further include one or more pharmaceutically acceptable carriers selected from the group consisting of fillers, extenders, binders, wetting agents, disintegrants, surfactants, diluents and excipients.

According to another aspect of the present invention, there is provided a food composition for antioxidant or cancer improvement comprising an extract of larva powder in the Dongae and the like.

According to another aspect of the present invention, there is provided an antioxidant cosmetic composition comprising an extract of larvae powder to the Dongae and the like.

According to another aspect of the present invention, fecal soil of larvae is provided on Dongae, etc. bred using a feed containing water parsley grown hydroponically with germanium water.

According to another aspect of the present invention, there is provided a fertilizer additive including the powder of larvae to the Dongae and the like.

According to another aspect of the present invention, there is provided a fertilizer additive including larva fecal soil to the Dongae and the like.

According to another aspect of the present invention, there is provided a feed additive including the powder of larvae to the Dongae and the like.

According to another aspect of the present invention, there is provided a feed additive including the feces of larvae to the Dongae and the like.

According to the present invention, it was found that the antioxidant and anticancer composition comprising the larva extract and fecal soil extract in Dongae et al. by adding germanium to the larvae in Dongae et al. has the ability to scavenging DPPH radicals and inhibiting the growth of cancer cells.

Therefore, the powder of larvae in Dongae, etc. bred using the parsley feed containing germanium of the present invention and the antioxidant and anticancer composition comprising the same can be used as a natural antioxidant or natural anticancer agent in medicine and pharmaceutical fields, and It can be usefully used as an antioxidant in the cosmetic field.

Figure 1 is a photograph showing the larvae (a) and fecal soil (b) in Dongae and the like containing germanium.
Figure 2 is a graph showing the DPPH radical activity for the ethanol extract of larva powder in Dongae et al. which does not contain germanium.
Figure 3 is a graph showing the DPPH radical activity for the ethanol extract of larva powder in Dongae et al. containing 0.42 ppm germanium.
Figure 4 is a graph showing the DPPH radical activity for the ethanol extract of fecal soil powder in Dongae et al. which does not contain germanium. Statistical analysis was performed using Duncan's multiple range test (p<0.05).
5 is a graph showing DPPH radical activity for ethanol extract of fecal soil powder in Dongae et al. including germanium. Statistical analysis was performed using Duncan's multiple range test (p<0.05).
6 is a graph showing the inhibitory effect of a larva extract containing germanium on the growth of LNCaP, a human prostate cancer cell line, after cell culture at 37° C. for 24 hours, 48 hours and 72 hours.
7 is a graph showing the inhibitory effect of a fecal soil extract containing germanium on the growth of LNCaP, a human prostate cancer cell line, after cell culture at 37° C. for 24 hours, 48 hours and 72 hours.

The present invention provides a powder of larvae (black soldier fly larva powder) to Dongae, etc. bred using a feed containing water parsley grown hydroponically with germanium water.

In one embodiment, the larvae on the Dongae etc. dry the water cultivated water parsley on average, and dry the water parsley on average 1 to 100 ppm, preferably 5 to 70 ppm, and most preferably 10 to 50 ppm of germanium water parsley: wet feed = 0.1 to 20: 100 (g/ml), preferably parsley: wet feed = 0.5 to 10: 100 (g/ml), most preferably parsley: wet feed = 1 to 5: 100 (g/ml) Included as, and a relative humidity of 0 to 40% general feed and a moisture control agent (for example, sawdust or rice husk, etc.) in a certain ratio (w/v) mixed feed (hereinafter referred to as "germanium compound feed" ) To breed.

Dongae, etc., which are bred with germanium compounded feed, can be directly reared using a conventional insect breeding machine, and can be provided with larvae raised with germanium compounded feed by consigning commercially.

Dongae et al. have a life cycle of 3 days from eggs, 15 days in larval stage, 14 days in pupa, 8-10 days for adults, and 40 to 42 days in total. By using the habit of moving to the place, you can easily separate the food and the dongae. In particular, it can be added to livestock feed using homeless larvae, which are at least 7 days after hatching and before pupa, which have the best nutritional status.

The larvae of Dongae, etc. bred with the germanium compounded feed of the present invention are placed in a cage of a certain size using an egg mass (algeb) from 24 to 36 ℃, preferably from 25 to 33 ℃, most preferably from 27 to 32 ℃ temperature, 50 Obtained by breeding for 5 to 10 days, preferably for 7 to 9 days, and most preferably for 8 days while maintaining a humidity of ~ 80%, preferably 55 ~ 75%, most preferably 60 ~ 70% Caterpillars (1000 ~ 15000% gain rate) can be harvested. Among them, caterpillars with a length of 25 ㎜ and a diameter of 7 ㎜ or more are selected and washed, and the loss of nutrients is reduced to a minimum and the moisture content is within 5 ~ 10%. have. In addition, the powder can be commercially provided from a company that entrusted the larvae to Dongae.

It can be seen that the germanium content of the larva powder in Dongae, etc. bred with the germanium compound feed is about 40 times higher than the germanium content of the larva powder reared with a general feed.

In addition, the present invention provides an extract of the larvae powder obtained by extracting the larvae powder to the Dongae and the like.

In one embodiment, the extract may be one or more selected from the group consisting of an ethanol extract, a methanol extract, a water extract, and a mixture thereof, preferably an ethanol extract.

The larvae of the powdered Dongae et al. may be extracted for a certain time at room temperature using ethanol as a solvent, and preferably 60 to 80% ethanol. In addition, the obtained larva extract can be stored under reduced pressure or freeze-dried.

In addition, the present invention provides a pharmaceutical composition for antioxidant or cancer treatment comprising an extract of larvae powder in the Dongae and the like.

Antioxidant activity was measured by reacting with DPPH (1,1-diphenyl-2-pictrylhhydrazyl) solution in germanium-containing larva extract, whereas the larval extract reared as general feed did not show antioxidant effect. You can see that it represents.

In addition, as a result of confirming the inhibition of the growth of cancer cells using the human prostate cancer cell line LNCaP in the germanium-containing larva extract, it can be confirmed that it shows a concentration-dependent and time-dependent growth inhibitory effect. In particular, when administered at a concentration of 75 mg/ml for 24 hours, it can be seen that the growth inhibitory effect of about 50% is exhibited.

In one embodiment, the pharmaceutical composition may further include one or more pharmaceutically acceptable carriers selected from the group consisting of fillers, extenders, binders, wetting agents, disintegrants, surfactants, diluents and excipients.

Pharmaceutically acceptable carriers can be used by mixing saline, sterile water, Ringer's solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol, and one or more of these ingredients, and if necessary, antioxidants, Other conventional additives, such as a buffer solution and a bacteriostatic agent, may be added.

In addition, the pharmaceutical composition of the present invention may be prepared in the form of an aqueous solution, suspension, emulsion, etc., in various dosage forms such as injection formulations, pills, capsules, granules, ointments, oral administrations or tablets.

In addition, the present invention provides a food composition for antioxidant or cancer improvement comprising a larva extract in the Dongae and the like.

The food composition of the present invention refers to a natural product or processed product containing one or more nutrients, and preferably refers to a state that can be eaten directly through a certain amount of processing process, and generally, health functional food , Beverages, food additives, and beverage additives may be included and used, but are not limited thereto.

In addition, the present invention provides an antioxidant cosmetic composition comprising a larva extract to the Dongae and the like.

The antioxidant and whitening cosmetic composition of the present invention may be used in basic cosmetics, makeup cosmetics, and body care cosmetics, but is not limited thereto. In addition, in the case of using the larvae extract in Dongae containing germanium according to the present invention as a cosmetic, purified water, oil, surfactant, moisturizing agent, higher alcohol, thickener, chelating agent, colorant, fatty acid according to the formulation or purpose of use of the cosmetic , Antioxidants, preservatives, waxes, pH adjusters, fragrances, etc. can be added to prepare a cosmetic composition.

In addition, the present invention provides fecal soil of larvae to Dongae, etc. bred using a feed containing water parsley grown hydroponically with germanium water.

Fecal taro on Dongae et al., produced in the process of growing larvae on Dongae et al., for example, is obtained by processing eggs, larvae or mixtures thereof, and decomposes organic waste (e.g., food waste, etc.) It may be a leftover product.

In addition, the present invention provides a fertilizer additive including the powder of larvae to the Dongae and the like.

In addition, the present invention provides a fertilizer additive including larva fecal soil to the Dongae and the like.

Recently, crops using germanium-added fertilizers are known to have the effect of preventing diseases and pests, and the growth of crops has increased after germanium is added, and the germanium-containing ore is finely pulverized, or the pulverized product is mixed with water to form a liquid and added to fertilizers. It is a trend to use. However, when germanium is added at a certain concentration or less, the effect hardly appears, and a sufficient amount of it has to be added, resulting in a high cost, so germanium is mainly used as a fertilizer additive for crops with a high unit cost.

On the other hand, the fecal soil in Dongae, etc. bred with the germanium compound feed can be confirmed to be significantly higher in germanium content (about 140 times or more) than the feces soil in Dongae, etc. raised as general feed, so that it is not only functional mushrooms and fruits and vegetables, but also weak to diseases and pests. It can be used conveniently as a differentiated fertilizer additive when growing crops.

In addition, compared to the fact that the antioxidative effect was not shown in the fecal soil raised with general feed, the result of measuring the antioxidant activity by reacting with the DPPH solution in the fecal soil containing germanium showed that it was lower than the larvae in Dongae, but showed statistically significant concentration-dependent DPPH scavenging activity. I can confirm.

In addition, the present invention provides a feed additive containing larva powder to the Dongae and the like.

In addition, the present invention provides a feed additive containing fecal soil to the Dongae and the like.

The feed additive may include ingredients commonly used in feed manufacturing, such as glucose, peptone, yeast extract, diphosphate, magnesium sulfate, and cysteine, but is not limited thereto.

Hereinafter, the present invention will be described in more detail through examples. However, the following examples are for illustrating the present invention, and the scope of the present invention is not limited thereto.

<Example>

1. Materials and methods

(1) Breeding in Dongae

Breeding in Dongae, etc. was requested by Entomo Co., Ltd. (formerly Korea Insect Resource Research Institute). After hydroponically cultivation of water parsley with germanium water, it was dried, and 200 ml of wet food and 200 ml of sawdust were added to 10 g of parsley containing 40 ppm germanium in the same ratio (v/v). 1a) and fecal soil (see FIG. 1b) were dried powder.

Dongae et al. larvae are 3 instar larvae obtained by breeding for 8 days while maintaining a temperature of 27 ~ 32 ℃ and humidity of 60 ~ 70% using about 0.5 g of egg mass in a 274 * 450 cm cage (about 25 Mm) 600 g (larvae gaining rate 1200%) was harvested.

After washing the larvae on Dongae etc. with a length of 25 ㎜ and a diameter of 7 ㎜ or more, the larvae were put in a dryer while the water was removed and dried with warm air at 50° C. for 36 hours. The larvae and fecal soil dried to a moisture content of 5 to 10% were pulverized in a grinder and powdered, mixed with 45 ml of 70% ethanol at a ratio of 5 g of powder, placed in a shaker, and sufficiently stirred for 24 hours. Antioxidant and anticancer efficacy were evaluated using the supernatant obtained by centrifugation.

(2) Cell culture and MTT assay

A culture medium containing 10% fetal bovine serum and penicillin-streptomycin in RPMI 1640 was used as a medium for the LNCaP cell line, a prostate cancer cell. Counting 3 × 10 ⁵ LNCaP cell lines into a 100 mm culture dish, and culturing in a 5% CO ₂ incubator at 37°C, black soldier fly larva powder extract (LPE) and black soldier fly larva powder extract (LPE) on Dongae, etc. Soldier fly feces soil extract, FSE) were treated at concentrations of 25, 50, 75, and 100 mg/ml, respectively, and 20 µl of MTT solution was added in a 96-well plate, followed by incubation for 4 hours. After centrifugation at 2,000 rpm for 10 minutes to remove the supernatant, 150 µl of DMSO was added to each well, stirred for 30 minutes, and absorbance was measured at 540 nm with a spectrometer.

(3) Antioxidant analysis

DPPH radical scavenging activity was measured as an antioxidant activity measurement method. 10 µl of a sample and 190 µl of a 0.2 mM DPPH (1,1-diphenyl-2-pictrylhhydrazyl) solution were diluted in a 96-well plate and reacted at room temperature for 30 minutes, and the absorbance was measured at a wavelength of 517 nm. DPPH radical scavenging activity ratio (Inhibition, %) was calculated as in Equation 1.

DPPH radical scavenging activity (%)=[(A _control -A _sample )/(A _control )]×100 <Equation 1>

2. Results

(1) Germanium content verification of larvae and fecal soil containing germanium

The germanium content of larvae and fecal soil raised with feed mixed with parsley was requested to be analyzed by the Korea Basic Science Institute. As shown in Table 1, 0.42 ppm of germanium was detected in larvae raised with parsley blended feed, 5.84 ppm of germanium was detected in fecal soil, 0.01 ppm of germanium was detected in larvae raised with general feed, and 0.04 ppm germanium was detected in fecal soil.

(2) Antioxidant effect of germanium-containing larva and fecal soil

The larvae and feces reared with normal feed as a control group did not show antioxidant effects (see FIGS. 2 and 4). In contrast, DPPH scavenging ability was shown in both germanium-containing larvae and fecal soil (see FIGS. 3 and 5), and in particular, fecal soil extract (FSE) having a high germanium content at a concentration of 50 mg/ml was 29.8%, and larva extract (LPE) was The DPPH scavenging ability of 54.13% was shown, indicating that the DPPH scavenging ability was not proportional to the germanium content. Compared to the fecal soil extract (FSE), the larva extract (LPE) having a relatively low germanium content showed very high antioxidant activity (see FIG. 3).

(3) Anti-cancer effect of germanium-containing larvae and fecal soil

Using LNCaP, a human prostate cancer cell line, larva extract (LPE) and fecal soil extract (FSE) were administered at a concentration of 25, 50, 75, 100 mg/ml for 24 hours, 48 hours and 72 hours. Dependent on the cell growth rate was significantly reduced. In particular, after administration of larva extract (LPE) at a concentration of 75 mg/ml for 24 hours, it showed a growth rate of 53.3%, whereas fecal soil extract (FSE) was 60%, and 54% at a concentration of 100 mg/ml. It was found that the growth rate of the larva extract (LPE) significantly inhibited the growth of cells than the fecal soil extract (FSE) (see FIGS. 6 and 7 ).

(3) Conclusion

Since the larvae of Dongae etc. contain high-quality amino acids, fatty acids, and omega components, they are being developed as food substitutes and biodiesel, and are currently consumed as feed for fish farming and food for companion animals, but there are still no studies on cosmetic materials. Can't see

According to the prior art [Lalander et al., 2016, Science of the Total Environment 565:279-286], three drugs (Carbamazepine, roxithromycin, trimethoprim) and two insecticides (azoxytrobin, propiconazole) were applied to larvae in Dongae et al. As a result, it was confirmed that the half-life was significantly shortened compared to the control group without accumulating any components, and it was reported that composting larvae in Dongae et al. could delay the spread of drugs and pesticides in the environment. According to another prior art [Lee, et al., 2016, J of The Korea Industry Association 10(6):451-459], parsley grown in soil containing germanium using germanium source water and concentrated water. As a result of analyzing the germanium content of and lettuce and observing the physiological activity effect according to the content, 51.8 mg/g of parsley (99.3 mg/g) and lettuce (5.6 mg/g) grown with natural germanium source water of 18.3 mg/g It was reported that the germanium content of parsley (545.3 mg/g) and lettuce (79.6 mg/g) grown in g germanium enriched water was higher, and the antioxidant and anticancer effects were higher in proportion to the germanium content.

In this study, it was confirmed that the germanium content was significantly increased in larvae and fecal soils reared by mixing 10 g of dried parsley powder (40 ppm) hydroponically grown for 45 days with 20 ppm of concentrated water in feed to Dongae etc. compared to the control group. In addition, the germanium content of the powder extract and fecal soil extract of the larvae containing germanium was higher in the fecal soil extract, but the larva extract showed significantly higher activity than the fecal soil extract in DPPH scavenging and cancer cell growth inhibition. (p<0.05). Larvae reared with the germanium-containing parsley blended feed of the present invention showed antioxidant and anti-cancer effects in proportion to the concentration, whereas fecal soil was found to be inversely proportional. Fecal soil produced in the process of larvae rearing is a product left after decomposing organic waste and contains more germanium than larvae, so it is believed that it can be used as a differentiated fertilizer when cultivating functional mushrooms and fruits. These results strongly suggest that larvae in Dongae and the like can be used as a life care material for functional foods, pharmaceuticals, and cosmetics.

(4) Summary

Dongae et al. are known as environmental cleaning insects that treat food waste, but they contain high-quality amino acids and fatty acids, and secrete antibiotics, showing the possibility of industrial use as a material for cosmetics, medicines and health functional foods. In this study, Dongae et al., raised by mixing water parsley hydroponic with germanium water into feed, succeeded in producing germanium-containing larvae and fecal soil. 0.42 ppm and 5.84 ppm of germanium were detected in the dry powder of the larva and fecal soil, respectively, and the general larva and fecal soil contained 0.01 ppm and 0.04 ppm germanium, respectively. As a result of analyzing the DPPH scavenging ability and the growth rate of cancer cells in these extracts, the larvae containing a large amount of germanium compared to the general larva showed excellent antioxidant and anti-cancer effects (p<0.05).

Currently, the fecal soil in Dongae, etc. is marketed as fertilizer, and the fecal soil in Dongae, which contains a large amount of germanium produced in the present invention, is a by-product of larvae in Dongae, etc. It is useful as a differentiated fertilizer for cultivation of functional mushrooms and fruits and vegetables. It is believed that it will greatly contribute to creating In addition, from these results, it can be confirmed that larvae in Dongae and the like are highly likely to be used as functional health materials for life care such as cosmetics, pharmaceuticals, and health functional foods.

Claims (14)

Powder of larvae on Dongae, etc. bred using feed containing water parsley grown hydroponically with germanium water. The powder of larvae of Dongae et al. according to claim 1, wherein the parsley is contained in the feed as parsley: wet feed = 0.1 to 20: 100 (g/ml). The powder of claim 1, wherein the larvae of the larvae are bred for 5 to 10 days. An extract of powdered larvae of larvae obtained by extracting the powder of larvae on the larvae of any one of claims 1 to 3. The extract of larvae powder according to claim 4, wherein the extract is at least one selected from the group consisting of ethanol extract, methanol extract, acetic acid extract, water extract, and mixtures thereof. A pharmaceutical composition for the treatment of cancer comprising an extract of larva powder in Dongae and the like of claim 4. The method of claim 6, wherein the pharmaceutical composition further comprises one or more pharmaceutically acceptable carriers selected from the group consisting of fillers, extenders, binders, wetting agents, disintegrants, surfactants, diluents and excipients. Pharmaceutical composition. Antioxidant or cancer-improving food composition comprising an extract of larva powder in Dongae, etc. of claim 4. Antioxidant cosmetic composition comprising an extract of larva powder in Dongae, etc. of claim 4. Fecal soil of larvae on Dongae, etc. bred using feed containing water parsley grown hydroponically with germanium water. A fertilizer additive containing the powder of larvae in Dongae, etc. of claim 1. A fertilizer additive containing larva fecal soil in Dongae, etc. of claim 10. A feed additive containing the powder of larvae in Dongae, etc. of claim 1. A feed additive containing fecal soil of larvae in Dongae, etc. of claim 10.

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