Transfer of substances with dietary and medicinal properties from post-mushroom cultivation biomass into insect bodies

Abstract

The invention discloses an insect with increased levels of proteins, chitin, elements, amino acids, vitamins and / or sterols. These insects have higher levels due to the inclusion of mushroom biomass in their diet. The mushroom biomass comprises any one or more of sporocarp, stipe and / or mycelium of mushrooms. Insects rich in biomolecules in mushroom biomass can be used directly, or as supplements, to increase the level of biomolecules in humans or other animals that may eat insects or improved forms thereof.

Description

Cross-reference to related applications

[0001] This application claims priority under 35 § 119(e) of U.S.C. No. 18 / 637,458, filed April 17, 2024, and U.S. Provisional Application No. 63 / 460,622, filed April 20, 2023, the entire contents of which are incorporated herein by reference. Technical Field

[0002] This invention relates to methods and products developed for transferring substances from biomass after mushroom cultivation into insects. (Coleoptera) Coleoptera Diptera ( Diptera ) and Orthoptera ( Orthoptera Insect larvae are particularly well-suited for this transfer, and the transfer can be best achieved by feeding insect larvae. The transfer of these substances should be achievable on an industrial scale, and the amount of these substances should significantly increase the content of these components in insects relative to control insects, resulting in a significant enrichment of the final product. Background Technology Industrial-scale insect rearing and breeding is one of the most dynamic emerging branches in agricultural production and the biotechnology industry. Insects are rich in protein, vitamins, elements, and all essential amino acids. They have a remarkable ability to convert low-energy feed from agricultural and food industry byproducts into high-quality protein and fat in a low-emission manner. Coupled with their short fattening period, insects are at the forefront of the restructured food chain.

[0003] By 2027, global mushroom production is projected to reach 24 million tons annually. (Source: https: / / www.marketdataforecast.com / market-reports / button-mushrooms-market, (see also https: / / www.fortunebusinessinsights.com / industry-reports / mushroom-market-100197) In Europe and North America, the main cultivated mushroom is the button mushroom (Agaricus bisporus). Agaricus bisporusProducing one kilogram of ready-to-eat mushrooms typically requires more than two kilograms of cultivation substrate. However, after cultivation, the SMS (sludge medium) or SMC (sludge compost) is completely covered by mycelium and contains most of the mushroom's residual nutrients. To date, no satisfactory mechanical or chemical method has been found to separate the mycelium from the substrate and utilize the rich components contained within it. Mycelium is often composted and used as soil, which not only wastes the specific substances contained in the mycelium but also increases the carbon footprint of mushroom cultivation.

[0004] The cultivation of tree-grown / woodland mushrooms is similar. They are cultivated by inoculating various sizes of mycelium bales, primarily composed of deciduous tree shavings, grains (such as wheat, corn, soybean hulls, sorghum, etc.) and their processed products (such as bran and even nut shells), hemp straw, coconut, vermiculite, and / or cardboard. The following varieties are mainly cultivated in this way: oyster mushrooms (… Pleurotus ostreatus , Pleurotus eryngii ),mushroom( Lentinula edodes ), tea tree mushroom ( Agrocybe aegerita ), Hericium erinaceus ( Hericium erinaceus ), Buna-no-Mushroom ( Hypsizygus tessulatus ) and enoki mushrooms ( Flammulina velutipes In this technique, used mycelial bags are waste completely covered by mycelium and must be discarded. Since mushroom harvesting is typically completed within an interval / stage, the amount present at harvest time is much greater (sometimes even more) than the amount in the lateral stream. Given that some dietary and medicinal active compounds in mushrooms are only found in the mycelium, such as erinacines in Hericium sp., any recovery achieved through transfer to insects would be advantageous, especially since it is highly efficient and inexpensive. Of every kilogram of mushrooms available for sale, 15-25% of the harvesting and processing waste consists of stipes. This biomass is characterized by its high nutrient content and, like the substrate, receives no management or protection.

[0005] Another type of waste in traditional mushroom production is water, which is obtained after rinsing the mushrooms and is also a byproduct of blanching them. This wastewater is often rich in protein, and the liquid within it is also rich in nutrients.

[0006] Another source of liquid biomass is obtained through fungal culture, where culture media / mediums of in vitro mycelial crops and Fusarium microfungi can be collected from fungal protein bioreactors. This fungal cultivation method is gaining popularity, and the scientific and industrial communities generally consider it suitable for edible and medicinal mushrooms or Fusarium (Fusarium) Fusariumvenenatu ) has a more sustainable future for the cultivation of foodstuffs. One strain of Fusarium (F. venenatum) is commercially used to produce the single cell protein mycoprotein Quorn. Fusarium venenatum

[0007] As fungi are adulterous organisms, they digest before they ingest, secreting digestive enzymes into the external environment to obtain nutrients, so the fluid after in vitro cultivation is extremely rich in nutrients, making it ideal for use in medicine.

[0008] On the other hand, in the mycoprotein bioreactor, after production of mycoprotein using Fusarium (F. venenatum) species (in a Quorn or airlift fermenter), the culture fluid / substrate is centrifuged to produce a "mycelial slurry" containing about 75% water. The resulting product is called mycoprotein and, after rapid cooling, is ready for use in food production in some embodiments. Fusarium venenatum In the biomass produced, the RNA content is reduced to make the product compliant with the necessary health standards. The method for reducing the RNA content is to subject the biomass to a thermal shock at a temperature of around 64-65°C and to place the biomass in a separate reactor with a stirrer for about 20-30 minutes. At this temperature, the RNA degrades into nucleotides or nucleic acid monomers and diffuses out of the cells. Unfortunately, at this temperature, other biomass components also break down, resulting in a loss of about 35-38% of the quality of the potentially useful product. After reducing the RNA content, the liquid in the reactor is heated to 90°C together with the biomass. The biomass is then centrifuged and cooled.

[0009] The effect of the RNA reduction step is that the RNA leaks through the cell membrane, resulting in a loss of up to or more than 30% of the biomass in the form of liquid biomass to the filtrate (i.e. the liquid discharged from the centrifuge) or side stream.

[0010] The composition of the filtrate is shown in Tables A and B below:

[0011] Table A Average amino acid composition of typical supernatant / side stream (grams per 100 grams of dry matter)

[0012] Table B Average typical values for the chemical composition of the supernatant / side stream

[0013] The total solids content of the filtrate / side stream is typically about 1.3 grams per 100 millilitres, so it is a dilute biomass containing the target biomolecules.

[0014] ​​Dehydration / extraction of the fermenter "waste" (called filtrate or side stream) can produce a 5'-nucleotides-rich, powdery component that can be used as a yeast-free flavor enhancer in food and has been proven to have umami effect / flavor.

[0015] The study also focused on how to partially use the percolate (water that has passed through the solids) for fermentation, thus reducing the wastewater load and water consumption and further improving the sustainability of the process.

[0016] However, both processes tend to be very expensive and energy-intensive, and since they require high processing temperatures, most of the pharmaceutical ingredients are lost due to degradation.

[0017] In contrast, the percolate usually does not need to be dehydrated / extracted to be used as a liquid component in insect feed, transferring components with both dietary and pharmaceutical properties into the insect body. This transfer is similar to that of solid biomass.

[0018] It is generally known that feeding insects with biomass produced by various mushrooms is a way of disposing of waste. However, to the best of the inventors' knowledge, prior to the present invention, it was not known to use the biomass produced by mushroom cultivation and processing as insect feed to improve the quality of the final product (in the form of insect meat and / or crude protein, fat, and chitin) and to transfer the substances contained in the mycelium into the insect protein by biotechnological means and to increase the content of these substances in the insect body. The present invention takes advantage of the high content of biomolecules in mushrooms and their effective uptake by insect larvae to enrich the final product of mushrooms. SUMMARY

[0019] The present invention relates to the transfer of substances from the biomass of cultivated mushrooms (particularly the common Agaricus bisporus) into the bodies of Coleoptera, Diptera, and Orthoptera insects. The present invention also relates to the rearing and breeding of these insects on an industrial scale to significantly increase the content of these components in the insect body. BRIEF DESCRIPTION OF DRAWINGS

[0020] Figure 1 A general scheme for the growth of insect larvae using mushroom biomass is shown. DETAILED DESCRIPTION

[0021] In an embodiment, the present invention relates to increasing the content of biological elements present in insects. Insects in the larval stage obtained according to the rearing and breeding process of the present invention include and have the following characteristics: the content of biological elements, especially macroelements (K, Ca) and microelements (Cu, Fe, Zn) in the insects is significantly increased after feeding the insects with feed supplemented with cultivated mushrooms, compared to insects fed with feed not containing cultivated mushrooms. These substances are essential for all higher organisms. The insects in the larval stage also contain a higher content of exogenous amino acids, such as phenylalanine and tryptophan. In an embodiment, the content of phenylalanine and L-tryptophan is higher, which makes them extremely valuable materials that can be used as dietary supplements in human food, pet food and feed for farm animals. The production techniques for different insect species often differ fundamentally, and a production technique cannot necessarily be transferred to other techniques.

[0022] For Coleoptera insects, the insects use "dry breeding": the breeding and rearing substrate, which is also the dry base feed in the form of larvae, consists of cereals, flour milled from these cereals, bran and all by-products of flour milling. In "dry farming", the water is usually supplied to the insects in the form of endogenous water contained in the plant components. Water is supplied endogenously in all foods, including but not limited to all vegetables, fruits and parts thereof, other by-products of the processing of agricultural foodstuffs, and plant-derived products in any form (e.g. pieces, pulp, gel or jelly).

[0023] From the beginning of the life cycle, the dry base mixture (usually in the form of cereal bran) is the substrate in which the adult insects lay their eggs. The eggs hatch in the dry base mixture, and the larvae are also fed on this mixture until processing. Since the dry base mixture is both the basic feed and the substrate that expands the living space of the insects, the insects eat this feed continuously throughout the fattening period, and the proportion of insect manure gradually replaces the dry base mixture feed. In the final stage of fattening, only the larvae to be processed and insect manure remain in the fattening container.

[0024] For Orthoptera insects, too, "dry" breeding and rearing methods are used. The only exception is the manure, which does not form the living space of the larvae and should be removed regularly. However, the rest of the technology, which is fed with dry base feed supplemented with wet fruit / vegetable pieces, pulp, gel or jelly or with water alone, remains the same.

[0025] Conversely, in the "wet" technology most commonly used in the fattening of Diptera insects, the larvae live in a moist environment, which is usually composed of vegetables, fruits, meat products or even manure and parts thereof, which, as in the case of dry technology, are both their living space and feed. These foods are in turn transformed into the metabolic by-products, i.e. manure.

[0026] The substrate resulting from the cultivation of Agaricus bisporus (SMS / SMC) and the processing waste in the form of stipes can be used in various stages of production of various insect species. The mushroom processing waste and the SMS can be used in dry and wet processes.

[0027] For feeding the insects, after the last harvest of mushrooms, the substrate can be heat-sterilized together with the mulch, usually heating the substrate to 80°C for 6-8 hours. At any stage of the sterilization process, the mulch can be mechanically separated from the substrate or can remain completely undetached. If the mulch is not detached, the addition of peat helps to maintain the proper humidity of the insect rearing substrate and enriches the feces. If detached, the mulch can be recycled. The substrate with or without mulch is mechanically broken down into a granular fraction with a particle size of between about 0.01 mm and 20 mm. The comminution can be done with any equipment. For example, the breaking down device can be one or more of an industrial comminuter, a flour mill or a blender. Then, and by mixing the comminuted material with the appropriate amount of water (usually in a ratio of 1:1-5) as required by the species, a homogeneous mixture is prepared and tested.

[0028] The order can also be reversed for process reasons, i.e. the material can be hydrated first and then ground to the desired particle size fraction.

[0029] The material can be supplemented with any liquid, such as water (including industrial water), fruit juice, or, in a variant, the liquid from the mushroom rinsing and / or blanching, and / or the mycelium liquid and / or the fungal protein liquid as an alternative.

[0030] The material that is suitable is the entire waste after the last cultivation stage, including the SMS, the mulch and the stipes, either untreated or sterilized. After grinding, this material usually does not need to be watered, as the stipes themselves contain a lot of endogenous moisture. This material is suitable for dry and wet processes in the form of a fresh pulp. The raw material prepared in this way can also be processed into a gel or a jelly by techniques that inhibit the process of pulp decomposition. Experimental results have shown that the very rapid decomposition of the mycelium, while not negatively affecting the fattening of the insects themselves, leads to a reduction in the content of certain components, such as protein amino acids. This reduction prevents their transfer. A slower decomposition process allows a higher content of components.

[0031] In any case, the prepared feed raw material should be applied according to the actual needs and the production process used.

[0032] The processing of the stipe from the stipe of the mushroom stem waste is similar to the mycelium derived product. The stipe is obtained during the mushroom harvesting process. As with the fruit and vegetable pulps, the stipe material should be chopped in various ways to produce a particle fraction between about 0.01 millimeter to 20 millimeters. Alternatively, sieving can be performed to obtain a particle size of about 0.1 to 20 millimeters, or a particle size of about 1 millimeter to 10 millimeters, or a particle size of about 2 millimeters to about 5 millimeters. Due to its high water content, in embodiments, this material can be thickened by the addition of food and / or feed thickeners or other feed ingredients such as corn meal, whole kernel ground corn, or by the addition of a feed supplement in the form of barley beer spent grain (dry or wet). To thicken, another fungus can also be used, namely feed yeast or yeast slurry, which is a byproduct of brewing beer, wine, or spirits.

[0033] The resulting material will typically undergo very rapid decomposition, even at low temperatures, such as refrigeration temperatures. This rapid decomposition will, to some extent, strip the processed mushroom of those constituents that need to be transferred. In an embodiment, to slow down the decomposition process, the stipe can be flash cooled immediately after harvesting at 2-4°C, using a variety of techniques including, but not limited to, cooling techniques as well as vacuum cooling by reduced pressure. The raw material obtained in the form of a pulp should be stored under low temperature conditions and fed to the insects as soon as possible after processing. As a variant, the raw material can be processed into a gel or a jelly using a preservative additive to extend its shelf life.

[0034] Since both the SMS and the stipe are waste / byproducts of mushroom cultivation, they can typically be mixed together. The raw material can also be used. Depending on the relative quantity of stipe, the resulting mixture will vary in terms of hydration. In an embodiment, after grinding and sieving the material to a particle size of about 0.10 millimeter to 20 millimeters, water can be added depending on the desired degree of hydration. In an embodiment, the amount of water added is such that the pulp obtains the appropriate physical and chemical parameters required.

[0035] The method of rearing and / or breeding insects described in the present invention includes and is characterized in that the material described in the present invention is used for each of the developmental stages of the insects. The material of the present invention can also be used in different techniques, it can be used as a feed, a feed additive / supplement, or a substrate. The material of the present invention can be used in combination with any other component or material. In an embodiment, it is used to transfer the constituents of the biomass of the mushroom to the insects after mushroom cultivation and processing. The insects will effectively process the biomass of the mushroom, thus making available to humans and other higher animals the constituents of the mushroom that were originally present and have dietary and medicinal value.

[0036] The following section sets forth some of the advantages of the present invention.

[0037] Advantages of the invention In embodiments, the present application provides an increased content of beneficial molecules and / or one or more enriched end-products in insect larvae. In a variant, one advantage of the present application is that certain components (such as magnesium, calcium, tryptophan, etc.) are increased in content. Since these components all have recommended daily intakes and are important for human health, any vector that makes these molecules more available at higher content is useful. For example, these molecules can sometimes be used for their therapeutic effects on humans or other animals.

[0038] End-products enriched in this way can have enhanced antioxidant, anti-aging, anti-inflammatory, regenerative, activating, neuroprotective and anti-depressive properties, enabling their widespread use in nutraceuticals, supplements for athletes and convalescents, pharmaceuticals, and potentially in advanced medicine and cosmetics.

[0039] Enriched insect proteins contain healthy and nutritional components that enable their widespread use in the production of the highest quality feed for farm animals and aquaculture products. They provide better raw materials for highly specialized pet foods as well as for veterinary nutritional products and supplements.

[0040] In addition, the increase in chitin and melanin content opens innovative application prospects in heavy industry, for example in biodegradable electronics or in energy storage.

[0041] Examples of embodiments of the invention I. In embodiments, an experiment was conducted to demonstrate the difference in the content of selected biological elements in T. castaneum larvae between individuals fed a traditional feed mixture and individuals fed a traditional feed mixture supplemented with mushroom biomass obtained after cultivation and processing of Agaricus bisporus. Alphitobius diaperinus ) larvae during the larval stage, this mushroom biomass being obtained after cultivation and processing of Agaricus bisporus. A. bisporus

[0042] The insects used in the various experiments were from a culture of insects present in the inventors' stock. The experiments were conducted throughout the rearing period of the larvae, i.e. the period after hatching of the eggs. The larvae were fed with the feed disclosed herein for 5 weeks, allowing them to grow. The experiments were repeated 8 times, with 100 rearing containers per group each time, and the various experiments showed no significant difference in terms of BWG (body weight gain), FI (food intake), FCR (food conversion ratio) or rearing duration. Thus, the differences seen herein are entirely due to the difference in food available to each group of larvae. The results reported herein are all averages.

[0043] Throughout the fattening period, the insects were in rearing containers with a bottom surface area of 0.24 m2 2 ​in polypropylene containers at a temperature of 30°C, a humidity of 60% and in the dark. The same number of eggs was used for each experiment. The 5-day incubation period was followed by a 5-week rearing period. The insects were fed with the feed mixture every 56 hours according to the needs. The insects were separated from the faeces and the mass of the insects was determined, and the food residues were weighed at the end of the cycle. No dead insects were found in all samples. The feed mixture used was as follows: A. Feed mixture, consisting of: - 35% (mass) wheat bran, 60% (mass) carrot pulp, 5% (mass) feed additive in the form of barley beer lees.

[0044] B. Compound feed, consisting of: - 35% (mass) wheat bran, 60% (mass) pulp of mushroom biomass after the last harvest (containing SMS residues and mushroom stipes), and 5% (mass) feed additive in the form of barley beer lees.

[0045] The pulp of the substrate and the mushroom stipes was prepared as described, with a degree of hydration of 70-90%, and was applied at intervals of 56 hours immediately after preparation. After the end of the rearing period, the larvae were separated from the faeces and the remaining unprocessed feed. The larvae were dried and ground to a powder. Samples were taken from the material prepared in this way and were then analysed.

[0046] The biomass obtained was homogenised in an agate mortar and subjected to mineralisation. The elemental composition of the samples was analysed using a Nanohunter II X-ray fluorescence spectrometer (TXRF) produced by the Japanese company Rigaku, which uses total reflection of X-ray radiation. The Nanohunter II instrument contains a 600 W molybdenum tube X-ray source and is equipped with a 16-sample loader, which ensures automatic operation of the device.

[0047] The results obtained confirm that in all cases the biological elements are transferred from the feed to the insect larvae. However, after the addition of the post-production residues of Agaricus bisporus, the concentration of almost all the required elements increased significantly (see Table 1).

[0048] Table 1 shows the content of selected elements in the insect larvae fed with traditional feed throughout the entire rearing period (insects A) and in the insect larvae fed with feed supplemented with biomass according to the development stage after mushroom cultivation (insects 1-4). The group listed as “insects 1” was reared for 2 / 3 of the rearing period, while the group listed as “insects 4” was reared for the entire rearing period. The results are expressed in milligrams of the respective element per kilogram of dry weight.

[0049]

[0050] N = 6, Se as internal standard Table 1 Analysis of the tables shows that almost all elements increased with the consumption of mushroom biomass by insects (cultivated and processed as described in this article). The analysis results show the percentage increase in the content of the following elements: Potassium content increased by 6.7%. Calcium content increased by 69%. Manganese content increased by 12%. Iron content increased by 37.6%. Copper content increased by 5.99%. Zinc content increased by 10.4%. Rubidium content increased by 66.6%. The results in Table 1 show that the transfer of macroelements (K, Ca) and microelements (Cu, Fe, Zn) was increased in the group fed with feed supplemented with mushroom biomass. These elements are well known to be essential for human health. Therefore, utilizing insect larvae containing higher levels of these elements should provide them with particular value in the diet. Notably, the tested material had a very high zinc content, making it easily usable as a dietary supplement to meet the daily human requirement for this element. Similarly, insect larvae with higher zinc content can serve as a good source of iron and also as a supplement to the human diet. The experimental results of both groups fed with mushroom biomass were superior to the control group in terms of these elements.

[0051] Larvae fed with mushroom biomass showed higher levels of most elements. Subsequent tests were conducted to determine whether insect larvae could obtain similar results from essential dietary organic molecules.

[0052] II. The aim of the next experiment was to determine the difference between individuals fed a conventional feed mixture and those fed with button mushrooms derived from cultivated and harvested processes. A. bisporus The biomass additives between individual small mealworms ( Alphitobius diaperinus Are there differences in the content of selected organic compounds in the larvae?

[0053] The insects used in the experiments were from the inventor's breeding inventory. Each experiment was conducted throughout the entire fattening period of the larvae, specifically the five weeks from egg hatching to larval maturity (and readiness for processing). The experiments were repeated eight times, with 100 rearing containers per group each time. There were no differences between groups in terms of BWG (body weight gain), FI (food intake), FCR (feed conversion ratio), or fattening time. Therefore, the differences in the relative levels of various biomolecules can be attributed to the different diets. The results have been averaged.

[0054] During the whole fattening period, the insects were kept in polypropylene containers with a bottom surface area of 0.24 m 2 at a temperature of 30°C, a humidity of 60% and in the dark. The same number of beetle eggs was used for each experiment. The 5-day incubation period was followed by a 5-week fattening period. The insects were fed every 56 hours according to the requirements. After the insects were separated from the faeces, the mass of the insects was determined and the food residues were weighed at the end of the period. No dead insects were found in any of the samples. The feed mixture used was as follows: Feed mixture, consisting of: A. - 35% (mass) wheat bran, 60% (mass) carrot pulp, 5% (mass) feed additive in the form of barley beer lees.

[0055] B. Compound feed, consisting of: - 35% (mass) wheat bran, 60% (mass) pulp of mushroom biomass after the last harvest (containing SMS residues and mushroom stems), and 5% (mass) feed additive in the form of barley beer lees.

[0056] The pulp of the substrate and the mushroom stems was prepared as described, with a degree of hydration of 70-90%, and was applied immediately after preparation at intervals of 56 hours. After the fattening period, the larvae were separated from the faeces and the remaining unprocessed feed. The larvae were dried and ground to a powder. Samples were taken from the material prepared in this way and were then analysed.

[0057] The insects were homogenised in a marvellous mortar. Samples of a suitable weight were selected by experiment and were extracted in an ultrasonic bath (Sonic-2, Polsonic) with methanol at a frequency of 40 kHz for 20 minutes. Each sample was repeatedly extracted several times with methanol. The repeatedly extracted juice was combined (300 mL) and concentrated. The evaporated extract was quantitatively dissolved in HPLC methanol and then filtered with a membrane filter (Millex, Millipore, USA). The extract thus obtained was ready for analysis by high-performance liquid chromatography.

[0058] L-tryptophan analysis L-tryptophan in the methanolic extract was determined by HPLC using a Hitachi-Merck (Dublin, Ireland) high performance liquid chromatograph. The apparatus equipped with an L-7100 pump, an L-7400 UV-VIS detector and a Purospher® (Dublin, Ireland) column was used to determine indole (e.g. tryptophan) compounds. The temperature of the Purospher® RP-18 column (4 mm x 200 mm, 5 pm) was 25 °C and the UV detection was recorded at l = 280 nm. The liquid phase used was a mixture of methanol / water / ammonium acetate in a volume ratio of (15:14:1 v / v / v) at a flow rate of 1 mL / min. The indole compounds were quantitatively analysed using the Beer’s law and a calibration curve assuming a linear relationship between absorbance and standard concentration. The results were expressed in milligrams per 100 grams of dry matter. An isocratic elution procedure was used to determine the indole compounds.

[0059] The results were calculated from the standard curve of L-tryptophan standard, i.e. the relationship between the area under the peak and the concentration of the compound.

[0060] Each test material was analysed in 6 independent replicates. The results (see Table 2) were presented as mean values and standard deviation (SD).

[0061] phenylalanine analysis The phenylalanine content was determined using an RP-HPLC method using a VWR Hitachi-Merck (Dublin, Ireland) high performance liquid chromatograph equipped with an L-2200 autosampler, an L-2130 pump, an RP-18e LiChrospher column (4 mm x 250 mm, 5 pm, thermostated at 25 °C), an L-2350 column detector and an L-2455 diode array detector. The column temperature was 25 °C. The mobile phase consisted of a mixture of solvent A: methanol and 0.5% acetic acid (volume ratio 1 :4) and solvent B: methanol. The gradient was: 0-25 min 100:0, 35 min 70:30, 45 min 50:50, 50-55 min 0:100, 57-67 min 100:0. The presence of phenylalanine in the analysed sample was identified by comparison of the UV spectrum and retention time with that of a standard (standard). The free phenylalanine was quantitatively analysed using a calibration curve assuming a linear relationship between absorbance and standard concentration (Table 2).

[0062] ergothioneine analysis The ergothioneine content was analyzed using high performance liquid chromatography (RP-HPLC) (Merck Hitachi, Tokyo, Japan), UV detector L-7400 (Merck Hitachi, Tokyo, Japan), pump L-7100, thermostat L-2350, chromatographic column RP-18 4 x 250 mm (Purospher®, particle size 5 μιη).

[0063] The analysis was performed using isocratic elution. A mixture of water and methanol in a volume ratio of 99:1 was used as mobile phase, to which 3.0 g of boric acid was added, in order to achieve a pH value of 5.0 for the solution. The flow rate of the mobile phase was set to 0.5 mL / min. 20 μί of sample were applied to the chromatographic column. The measurement was performed at a wavelength of 257 nm for 20 minutes. Ergothioneine was quantitatively analyzed using a calibration curve, assuming a linear relationship between absorbance and standard concentration (see Table 2).

[0064] Table 2 shows the content of selected organic compounds in insect larvae fed with traditional feed (insects A) and in insect larvae fed with feed supplemented with biomass after mushroom cultivation, depending on the developmental stage. The group listed as insects 1 is larvae in the 2 / 3 of the fattening period, the group listed as insects 4 is larvae analyzed after the end of the entire fattening period. The content of the various organic compounds is expressed in milligrams of biomolecule per 100 grams of dry weight of insect larvae.

[0065]

[0066] N = 6 Table 2 The analysis results show that the content of selected amino acids is significantly increased (up to more than 450%) in insects fed with mushroom cultivation and processing biomass as feed compared to insects fed with traditional feed mixtures. It is particularly noteworthy that the amino acid content reaches a peak in the 2 / 3 of the fattening period (after which it begins to decrease). Without being bound by theory, it is believed that this result is due to the fact that in the last fattening period, the larvae use the accumulated amino acids to prepare for the transformation into adult form.

[0067] In embodiments, insect larvae fed with mushroom biomass as feed can produce higher levels of essential amino acids. Essential amino acids include histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine.

[0068] Subsequently, experiments were performed to determine the relative content of ergosterol in control groups of larvae and larvae fed with feed supplemented with mushroom biomass.

[0069] ergosterol Table 3 shows the results of ergosterol content in larval insects fed with mushroom- added feed compared to the control group (i.e. larval insects fed with feed not added with mushrooms). The preparation of the groups was as described above. The groups listed as insects 1 were larvae grown for 2 / 3 of the five-week growth period, and the groups listed as insects 2 were larvae grown for the full five-week growth period.

[0070] Ergosterol content in insect samples is expressed in milligrams of ergosterol per 100 grams of dry matter (dm)

[0071] Table 3 From the results of Table 3, it can be seen that the ergosterol levels in the larvae fed with feed added with mushroom biomass were higher compared to the control group, which indicates that the ergosterol levels in the larvae increase when they are fed with feed added with mushroom biomass.

[0072] Five grams of material were extracted with a mixture of methanol / dichloromethane 75:25 (v / v). The mixture was sonicated (40 kHz) for 10 minutes. After 2 hours, the extract was centrifuged at 12,000 rpm and the supernatant was decanted. The extraction process was repeated twice and the resulting extracts were combined and evaporated to dryness. The determination was performed using high-performance liquid chromatography-diode array detector (HPLC-DAD). The determination was performed using a VWR / Hitachi LaChrom Elite liquid chromatography kit (Merck Hitachi, Tokyo, Japan) and a DAD L-2455 detector. An RP-18 4 x 250 mm (LiChrosfer) column was used with a particle size of 5 pm, a thermostat L-2350, a pump L-2130, and an automatic sampler L-2200.

[0073] The mobile phase was composed of solvent A: methanol / water 80:20 (v / v) and solvent B: methanol / dichloromethane 75:25. The gradient program was: 0-10 minutes, 80:20% B; 10-35 minutes, 40-60% B; 35-50 minutes, 0-100% B; 50-55 minutes, 80-20% B; hold at 25 °C for 15 minutes). The flow rate was 1.0 mL / min. The chromatographic peaks were recorded at 280 nm. The sterol standard was purchased from Fluka (Chemie AG).

[0074] Subsequently, various vitamins tests were performed on the larval groups to determine whether the larvae fed with feed added with mushroom biomass exhibited higher vitamin levels than the control group (i.e. larvae fed with feed not added with mushroom biomass).

[0075] Table 4 shows the content of selected vitamins in insect larvae fed with traditional feed (A) and in insect larvae fed with feed supplemented with biomass after mushroom cultivation, depending on the growth phase, throughout the rearing period. Group 01 is 2 / 3 of the rearing period, group 04 is the entire rearing period. Vitamin content is expressed in micrograms of vitamin per 100 grams of dry weight.

[0076]

[0077] Table 4 As can be seen from Table 4, the addition of mushroom biomass to the feed of the larval insects greatly and unexpectedly increased the content of thiamine (B1), riboflavin (B2) and nicotinic acid, compared to the control group.

[0078] thiamine (B1) and riboflavin (B2) The content of thiamine (B1) and riboflavin (B2) was determined using high-performance liquid chromatography. The high-performance liquid chromatography samples were prepared according to the method of PN-EN 14122:2004 / AC:2006 and PN-EN 14152:2004 / AC:2006. Thiamine and riboflavin were determined after an oxidation reaction before the chromatographic column. For this purpose, a 0.04% solution of potassium hexacyanoferrate (III) (dissolved in a 15% solution of sodium hydroxide) was added to the sample, which was shaken and left to stand for 2 minutes. After adjusting the pH to 7 with a phosphoric acid solution, it was centrifuged, the extract was purified by solid-phase extraction (SPE), and after a new centrifugation, it was subjected to high-performance liquid chromatography (HPLC) analysis. The Merck Hitachi high-performance liquid chromatograph (HPLC) equipped with an L-7612 online degasser, an L-7250 programmable autosampler, an L-7100 pump, an L-7480 fluorescence detector, an L-7360 column oven and a D-type interface was used for vitamin detection. The software detection program used was: D-7000 HPLC - System - Manager (HSM). The analysis was carried out on a Bionacom Velocity C18 PLX 4.6 x 250 mm, 5 μm column produced by Bionacom LTD (UK) and a pre-column produced by the same company. The measurement was carried out at an excitation wavelength and emission wavelength of 360 / 503, so that thiamine and riboflavin could be determined simultaneously. The mobile phase was water and acetonitrile. Gradient elution was used: t = 0 w / ac 88 / 12; t = 12 w / ac 0 / 100, at a temperature of 22°C. Thiamine and riboflavin were identified and quantified using thiamine and riboflavin external standards in hydrochloric acid and acetic acid, respectively.

[0079] Nicotinic acid (B3) The vitamin B3 content was determined using a Merck HITACHI high-performance liquid chromatograph (HPLC) with the improved method described by Juraj et al. (2003).

[0080] III. The purpose of the next experiment is to demonstrate that, after the cultivation and harvesting process, the button mushroom (… A. bisporus Between individuals fed a feed mixture in the form of substrate residue and stipes from the last crop of mushrooms, and individuals fed only with post-cultivation substrate (SMS) without any mushroom casing soil and stipe residue, small mealworms ( Alphitobius diaperinus Differences in the content of selected elements, organic compounds, vitamins and sterols in the larvae.

[0081] The insects used in the experiment were obtained through artificial rearing. Each experiment was conducted throughout the entire fattening period of the larvae, from egg hatching to larval maturity after 5 weeks. The experiment was repeated 8 times, with 100 rearing containers used per group each time. They did not show any differences in body weight gain (BWG), food intake (FI), food conversion ratio (FCR), or fattening time. The results are averaged.

[0082] Throughout the fattening period, the insects had a bottom area of ​​0.24 m². 2 The insects were reared in polypropylene containers at 31°C and 60% humidity in complete darkness. The same number of beetle eggs were used in each experiment. After a 5-day incubation period, they entered a 5-week fattening period. Feed was provided every 56 hours as needed. At the end of the life cycle, the insects were separated from their feces and food residue and weighed to determine their weight. No dead insects were found in any samples. The feed mixture used was as follows: The feed mixture has the following composition: -35% (by weight) wheat bran, 60% (by weight) post-harvest slurry of mushroom biomass (including SMS residue and mushroom stems), 5% (by weight) feed additives in the form of waste barley grains. B. Feed mixture, with the following composition: -35% (by weight) wheat bran, 60% (by weight) biomass slurry from clean SMS after cultivation (without any stipes or casing soil), and 5% (by weight) feed additives in the form of barley brewer's grains.

[0083] The preparation method of the substrate slurry is as follows: the raw substrate from the last crop is mechanically separated from the casing soil and mushroom stems. Then, water is added to the substrate at a weight ratio of 1:1.5, and the resulting mixture is ground into uniform particles, as described in the granulation process. The slurry is applied immediately after preparation at 56-hour intervals.

[0084] After the entire fattening period, the larvae are separated from their feces and uneaten feed residue.

[0085] The larvae were dried and ground into powder. Samples were obtained from the material prepared by this method and analyzed using the previously described method.

[0086] The selected components were detected in the diets of insects fed feed mixture B. These results are shown in Tables C, D, and E.

[0087]

[0088] Table C

[0089] Table D

[0090] Table E The analysis showed that the vitamin content was relatively high, the ergosterol content was low, the content of certain amino acids was high, and the element content was significantly low. This is understandable considering the slurry dilution ratio was 1:1.5, but it also indicates that the mycelium itself contains an exceptionally high concentration of B vitamins and the tested amino acids.

[0091] IV. The aim of the next experiment was to determine the difference between individuals fed a conventional feed mixture and those fed with button mushrooms derived from the harvesting process. A. bisporus The stem of the small mealybug is a biomass additive in the form of a small individual. Alphitobius diaperinus Is there a difference in chitin content in the larvae?

[0092] The insects used in the experiments were from the inventor's stock of farmed insects. Each experiment was conducted throughout the entire fattening period of the larvae, specifically the five weeks from egg hatching to larval maturity (and readiness for processing). The experiments were repeated eight times, with 100 rearing containers per group each time. There were no differences between groups in terms of BWG (body weight gain), FI (food intake), FCR (feed conversion ratio), or fattening time. Therefore, the differences in the relative levels of various biomolecules can be attributed to the different diets. The results have been averaged.

[0093] Throughout the fattening period, the insects had a bottom surface area of ​​0.24 m². 2The insects were kept in polypropylene containers at a temperature of 30°C, a humidity of 60% and in the dark. The same number of beetle eggs was used as starting material for each group. The 5-day incubation period was followed by a 5-week fattening period. The insects were supplied with feed every 56 hours according to requirements. The mass of the insects was determined after separating the insects from their faeces and food residues, and the weight of the larvae was weighed at the end of the cycle. No dead insects were found in any of the samples. The feed mixture used was as follows, as described above: Feed mixture, consisting of: A. - 35% (by mass) wheat bran, 60% (by mass) carrot pulp, and 5% (by mass) feed additive in the form of barley beer lees.

[0094] B. Compound feed, consisting of: - 35% (by mass) wheat bran, 60% (by mass) mushroom biomass pulp (refers only to the stipe after the last harvest), 5% (by mass) feed additive in the form of barley beer lees.

[0095] The pulp of the mushroom stipe was prepared according to the described method and the feed was applied at intervals of 56 hours immediately after preparation. After the fattening period, the larvae were separated from the faeces and the remaining unprocessed feed. The larvae were dried in the sun and ground to a powder. Samples were taken from the material prepared in this way and then analysed.

[0096] The chitin content test was carried out according to the methodology set out by the Forschungsinstitut Futtermitteltechnik der IFF, Frickenmuhle 38110 Braunschweig-Thune, Germany, as described in the following references, the entire contents of which are hereby incorporated by reference: https: / / www.researchgate.net / publication / 373317761_Chitin_in_Futtermitteln_-_Analyse_Auswirkung_auf_den_Nahrwert_und_Kontrolle https: / / www.researchgate.net / publication / 369977223_Bestimmung_des_Chitingehaltes_von_insektenbasierten_Produkten_Insekten_als_Proteintrager_Chitin_als_Nahrstoff https: / / www.researchgate.net / publication / 363491517_Bestimmung_des_Chitingehaltes_von_insektenbasierten_Produkten In short, the chitin content can be determined by following these steps.

[0097] The sample was ground to a particle size of less than approximately 1 mm.

[0098] Weigh 1.00 gram of sample and place it in a fiber bag.

[0099] The sample was degreased (oil removed) using petroleum ether.

[0100] Alkaline hydrolysis was carried out at 100°C for 60 minutes with 0.25 M NaOH.

[0101] Wash the alkaline hydrolysis products twice with hot water to remove them.

[0102] Nitrogen content can be determined using the Kjeldahl method. In short, the process involves three steps: 1) digestion; 2) distillation; and 3) titration. In the presence of a catalyst (such as selenium) and sulfuric acid, nitrogen species are converted to ammonia through wet combustion, with the ammonia remaining as ammonium sulfate. The ammonium sulfate is treated with NaOH to produce ammonia, which is then distilled to boric acid. The resulting borate anion is titrated with HCl to determine the nitrogen content.

[0103] Chitin content can be determined using the following formula.

[0104] Chitin (%) = ((V - Vblind(ml) * 100 * 1.4007 / msample(mg) * 14.007(g / mol N)) / (203.19(g / mol GlcNAc))) The results unexpectedly showed that insects fed mixture B not only had significantly increased chitin content, but also increased protein content and decreased fat content (see Table 5).

[0105] Table 5: The basic protein and fat composition of all samples were analyzed according to the LUFA method.

[0106] V. To further illustrate the performance, an experiment was conducted to demonstrate the effectiveness of feeding black soldier flies with a feed mixture containing mushroom biomass additives. Hermetia illucens The melanin content in the larval stage increases; this additive is produced through the cultivation and processing of button mushrooms. A. bisporus It was obtained after that.

[0107] The reason for conducting these tests was that the various larvae fed with mushroom biomass showed a significant color change compared to those fed a mixture without mushrooms. Depending on their developmental stage, black soldier fly larvae are light brown, gradually turning dark brown in the pre-pupal stage. According to the inventors' observations, it was clear that larvae fed a feed mixture containing fungal biomass immediately turned dark brown, almost black in the pre-pupal stage. Individuals at this developmental stage are closer to the adult—the adult insect.

[0108] The experiment was conducted throughout the entire rearing period of the larvae, that is, during the period after egg hatching. The larvae were fed the diet disclosed herein for 14 days (including the 3-day hatching period).

[0109] Throughout the fattening period, the insects had a bottom surface area of ​​0.64 m². 2 They were reared in polypropylene containers at a temperature of 32-34°C and a humidity of 75-85%. The same number of eggs were used in each experiment. This perishable feed additive, containing mushroom stalks and slaughterhouse waste, was given in slurry form every 48 hours as needed.

[0110] The experiment was repeated three times, with 10 rearing containers used per group each time. After the cycle, the insects were separated from feces and food scraps and weighed. No dead insects were found in any samples. The feed mixture used was as follows: A. Feed mixture, with the following composition: -75% crushed and hydrated wheat grains and wheat bran, 20% (by weight) feed additives in the form of barley brewer's grains, and 5% slurry of slaughterhouse waste (especially offal, head, paws and / or poultry carcasses).

[0111] B. Feed mixture, with the following composition: -75% is chopped and hydrated mushroom biomass (including mycelium and mushroom stems) after the last harvest, 20% (by weight) is feed additive in the form of barley brewer's grains, and 5% is slurry of slaughterhouse waste (especially viscera, head, claws and / or poultry carcasses).

[0112] At the end of the fattening period, the pre-pupae larvae are separated from their feces and remaining unprocessed feed. The larvae are then dried.

[0113] The test material was micronized / pulverized to a particle size of 3-4 mm. Then, 10% NaOH was added to the powder in a mass ratio of 1 : 10 and stirred repeatedly for 1.5 hours at 80°C in a closed container to isolate the chitin. Then, after cooling, an equal amount of 50% NaOH was added to the same container and a deacylation reaction was carried out at 95°C for 2.0 hours in a closed container with constant stirring. The resulting hydrolysate was filtered using a Buchner funnel. The residue (chitosan) was washed at pH = 7 until the wash water was neutral. Concentrated hydrochloric acid was used to precipitate the melanin at pH 2 from the alkaline hydrolysate, and then centrifuged at 1500 g for 15 minutes. The separated residue was washed to a neutral pH value. The residue was dried at 50°C to a constant weight (until no more weight was lost), or it was added to the previously isolated chitosan to obtain a melanin-chitosan complex, which was also dried at 50°C. When the melanin residue was centrifuged, it was separated into two parts - the actual residue and the hydrophobic part on the supernatant (i.e. the melanin layer and the fat complex layer). After separating the residue, the melanin and fat complex were separated using a separatory funnel. The separated components were dried as described above. The pigments were identified on the basis of their solubility, spectral properties and paramagnetic properties. Then, the dried samples were subjected to electron paramagnetic resonance (EPR) spectroscopy measurements. The resulting spectra were compared with standard spectra to determine the various components.

[0114] The lipids were extracted from the melanin-fat complex in a glass ampoule with a screw cap by adding 3 ml of a dichloromethane-methanol mixture (volume 10:1) and treating in an ultrasonic bath for 15 minutes. Then, the specific parameters of the gas chromatograph and the parameters of the mass spectrum recording are described.

[0115] A melanin-like pigment was isolated from two samples of black soldier fly larvae. The EPR signal g-factor of the sample A larval pigment was 2.0036 and the ΔΗ value was 6.3 ± 0.7 Gs. It was assumed that the pigment of the sample A larvae belongs to the eumelanin class. The concentration of the paramagnetic centers in the sample was 1.1 × 10 16 spin / g dry weight, which corresponds to 5.7 mg of eumelanin per 1 g of larva dry weight.

[0116] The melanin in the sample of insects fed on the feed mixture B was: ΔΗ = 5.7 Gs, g-factor = 2.0036, spin concentration 8.0 × 10 16 spin / g spin / g, which corresponds to a eumelanin content of 22 mg of eumelanin / g DM.

[0117] Larvae fed a biomass mixture containing the stipe and mycelium of *Agaricus bisporus* exhibited a nearly fourfold increase in eumelanin content, demonstrating the feasibility of using this method to obtain eumelanin on an industrial scale. In almost all cases, the presence of mushroom biomass supplementing the diet of various larval insect populations showed higher levels of elements, amino acids, vitamins, and / or steroids compared to the control group fed a diet without mushroom biomass. Insects are able to process and store additional elements and biomolecules, enabling their presence within the developing larvae.

[0118] Therefore, in the embodiments, the present invention relates to feeding experiments on various insects used in industrial production, the feed being by-products of mushroom cultivation, namely, substrates after mushroom (button mushroom) cultivation and processing waste.

[0119] In the implementation scheme, the present invention demonstrates unexpectedly excellent results and shows that desired substances with dietary and medicinal properties can be recovered from biomass after insect farming. By processing mushrooms, not only can these desired substances be biotransferred into the insects, but their effects can also be enhanced.

[0120] In its implementation, the present invention demonstrates that by adding mushroom biomass to insect feed, insect meat with previously unknown quality parameters can be obtained, with a quality far exceeding any product obtained from conventional fattening of conventional insect larvae using conventional feed ingredients.

[0121] In one embodiment, the present invention relates to insect larvae fed with a feed containing mushroom biomass, wherein the insect larvae contain higher levels of one or more elements, amino acids, vitamins and / or sterols compared to insect larvae fed with a feed not containing mushroom biomass.

[0122] In the embodiments, the mushrooms used in this invention are edible mushrooms. Edible mushrooms that can be used in this invention include, but are not limited to, white mushrooms (…). Agaricus bisporus ), brown mushroom (Italian brown mushroom), portobello mushroom, shiitake mushroom (forest shiitake or oak shiitake), oyster mushroom, porcini mushroom, morel mushroom, enoki mushroom (snow velvet mushroom), chanterelle mushroom (Girol mushroom) and / or maitake mushroom.

[0123] In one variant, the insect larvae are Coleoptera, Diptera, and Orthoptera. In another variant, the Coleoptera, Diptera, and Orthoptera are small mealybugs (…). Alphitobius diaperinus ), small fungi ( Alphitobius laevigatus ), yellow mealworms Tenebrio molitor ), black mealworms ( Tenebrio obscurus ), dark mealworm ( Tenebrio opacus), super mealworm (Zophobas atratus), super mealworm (Zophobas morio), black soldier fly ( Hermetia illucens ), houseflies ( Musca domestica ), house cricket ( Acheta domestica ), Short-winged hearth cricket (Gryllodes sigillatus), American cricket (Gryllus assimilis), Two-spotted cricket ( Gryllus bimaculatus ), East Asian migratory locust ( Locusta migratoria ), desert locusts Schistocerca gregaria ).

[0124] In one variant, the mushroom biomass is derived from the button mushroom (Agaricus bisporus). Agaricus bisporus In the implementation scheme, the mushroom biomass is derived from mushrooms, mycelium, and stipes. In one variant, these elements are one or more of potassium, calcium, manganese, iron, copper, zinc, or rubidium. In one variant, these amino acids are essential amino acids. In one variant, the amino acid is tryptophan, phenylalanine, or ergothioneine. In one variant, the vitamin is vitamin B1, vitamin B2, or niacin. In one variant, the sterol is ergosterol.

[0125] In an embodiment, the present invention relates to a method for increasing the levels of elements, amino acids, vitamins, and / or sterols in insect larvae and / or other animals or humans, the method comprising: Obtaining insect eggs, To cause the eggs to hatch into insect larvae, and Insect larvae were cultured in the presence of feed containing mushroom biomass.

[0126] In variations of the method, the insect larvae are Coleoptera and / or Diptera. and / or Orthoptera insects. In one variant, the mushroom biomass is derived from the button mushroom (Agaricus bisporus). In another variant, the mushroom biomass is derived from the mushroom, mycelium, and stipe. In one variant, these elements are one or more of potassium, calcium, manganese, iron, copper, zinc, or rubidium. In one variant, these amino acids are essential amino acids. In one variant, the amino acid is tryptophan, phenylalanine, or ergothioneine. In one variant, the vitamin is vitamin B1, vitamin B2, or niacin. In one variant, the sterol is ergosterol.

[0127] In one embodiment, the present invention relates to a method for transferring higher concentrations of elements, amino acids, vitamins, and / or sterols from mushrooms into insect larvae, the method comprising feeding the insect larvae with a feed containing mushroom biomass. In a variation of the method, the insect larvae are Coleoptera and / or Diptera and / or Orthoptera insects that feed on mushroom biomass derived from Agaricus bisporus. In another variation of the method, the elements are one or more of potassium, calcium, manganese, iron, copper, zinc, or rubidium; the amino acids are essential amino acids; the vitamins are one or more of vitamin B1, vitamin B2, or niacin; and the sterols are ergosterol. In one variation, the amino acid is one or more of tryptophan, phenylalanine, and / or ergothioneine.

[0128] The species listed below are all mushrooms of industrial importance, cultivated for food and medical purposes. Their mycelium, stipes, processing waste, and post-cultivation substrates can be used to transfer ingredients with dietary and medicinal properties into insects.

[0129] In one embodiment, the present invention relates to a method for increasing the chitin and protein levels of insect larvae, the method comprising feeding the insect larvae with a feed containing mushroom biomass.

[0130] In the embodiments, the mushrooms that can be used in this invention include Ganoderma lucidum, shiitake mushrooms and / or maitake mushrooms.

[0131] In the implementation plan, the mushrooms that can be used include oyster mushrooms (Pleurotus ostreatus). Pleurotus ostreatus (Jacq.) P.Kumm.), Yellow elm mushroom ( Pleurotus citrinopileatus (Singer), Florida Pleurotus ostreatus ( Pleurotus florida (Singer), lung-shaped lateral ear ( Pleurotus pulmonarius ((Fr.) Quél.), King Oyster Mushroom ( Pleurotus eryngii ((DC.) Quél.), Red oyster mushroom ( Pleurotus djamor (Rumph. Ex Fr.)Boedijn), Tea Tree Mushroom ( Agrocybe aegerita ), Elm-shaped pleated umbrella ( Hypsizygus ulmarius (Bull.)Redhead, Chicken Leg Mushroom ( Coprinus comatus ((OF Müll.) Pers.), Ganoderma lucidum ( Ganoderma lucidum (Curtis) P. Karst, Cordyceps militaris ( Cordyceps militaris (L.) Link), Cordyceps sinensis ( Cordyceps sinensis ), Ochre-scaled mushroom ( Agaricus subrufescens Peck, Giant King Mushroom ( Stropharia rugosoannulata) Farl. ex Murrill, Flammulina velutipes ( Flammulina velutipes ) (Curtis) Singer, Grifola frondosa ( Hypsizygus tessellatus ) (Buli.) Singer, Agaricus blazei ( Hypsizygus marmoreus ) (Buli.) Singer (Buna shimeji), Populus versicola ( Cyclocybe aegerita )( cylindracea ) (V. Brig.) Vizzini, Sparassis spathulata ( Sparassis crispa ) (Wulfen) Fr., Hericium coralloides ( Hericium coralloides ) (Scop.) Pers., Hericium erinaceus ( Hericium erinaceus ) (Buli.) Persoon, Irpex lacteus ( Tuber borchii ) Vittad., Indocybe indica ( Tuber indicum ), Polyporus umbellosus ( Polyporus umbellatus ) (Pers.) Fr., Grifola frondosa ( Grifola frondosa ) (Dicks.) Gray, Lentinula edodes ( Laetiporus sulphureus ) (Bull.) Murrill., Ganoderma lucidum ( Ganoderma lucidum ) (Curtis) P. Karst, Pholiota nameko (T. Itô) S. Ito et S. Imai., Coprinus comatus ( Kuehneromyces mutabilis ) (Schaeff.) Singer et A. H. Sm., Volvariella volvacea ( Volvariella volvacea ) (Bull.) Singer, Hericium coralloides ( Hericium coralloides ) (Scop.) Pers., Hericium erinaceus ( Hericium erinaceus ) (Buli.) Persoon, Lentinula edodes ( Lentinula edode s) (Berk.) Pegler (Shiitake), Auricularia polytricha ( Auricularia polytricha ) (Mont.) Sacc (mun mushroom), Grifola frondosa ( Grifola frondosa ) (Dicks.) Gray, Volvariella bombycina ( Psilocybe cyanescens ), Agrocybe aegerita ( Psilocybe cubensis ), Conocybe apala ( Psilocybe semilanceata ), Psathyrella mexicana ( Psilocybe mexicana ), Hypholoma sphaeriodes ( Psilocybe muscorumi ), Agrocybe aegerita ( Stropharia cubensis ) or Fusarium ( Fusarium sp. ) In embodiments, the target biomolecules transferred from the listed mushrooms into the insect body are: 5-hydroxytryptophan, L-tryptophan, serotonin, melatonin, ecdysone, ergosterol, lovastatin, ergothioneine, gamma-aminobutyric acid (GABA), phenolic compounds, melanin, cordycepin, erinacin, hericenones, enzymes, B vitamins, and macro and trace elements.

[0132] In embodiments, the present application relates to a feed for feeding insect larvae, the feed comprising a feed and a mushroom biomass. In one variant, the feed consists of wheat bran, post-cultivation mushroom biomass pulp, mushroom stipe, and feed additives. In one variant, the feed comprises about 35% (by weight) wheat bran, 60% (by weight) post-cultivation mushroom biomass pulp and mushroom stipe, and 5% (by mass) feed additives, wherein the feed supplements can be beer lees and / or meat by-products, which can be used as protein supplements.

[0133] In embodiments, the feed can comprise 20-70% (by weight) wheat bran, 20-75% mushroom biomass, and 1-25% (by weight) of feed additives such as barley lees. In one variant, the feed can contain 30-40% (by weight) wheat bran, 50-70% mushroom biomass, and 5-25% (by weight) of protein feed supplements such as barley lees and / or meat by-products.

[0134] In embodiments, the present application relates to a feed mixture, which can optionally contain beer lees and contains 1-25% of feed yeast and / or yeast cake. After the completion of brewing, a semi-liquid substance similar to yeast paste is formed at the bottom of the beer brewing tank, which is the yeast cake. This feed mixture can be used both in the larval rearing stage and in the reproductive adult stage.

[0135] The following references, as well as any reference cited herein, are incorporated by reference in their entirety for all purposes.

[0136] Juraja SM, Trenerry VC, Millar RG, Sheelings P, Buick DR. Asia-Pacific Food Analysis Network (APFAN) Training Exercise: Determination of Niacin in Cereals by Alkaline Extraction and High Performance Liquid Chromatography. Journal of Food Composition and Analysis (2003), 16, 93-106. doi:10.1016 / S0889-1575(02)00131-X.

[0137] It should be appreciated that, and it is also contemplated by the present application and within the scope of the present application, and within the scope of the present application, any of the above-listed features can be combined with any other of the above-listed features, as long as these features do not conflict with each other. When referring to a range, any real number meeting the range is considered to generate the end points of a sub-range. In summary, the present application is defined by the following claims.

Claims

1. Enriched insects fed a diet containing mushroom biomass, wherein the enriched insects contain elevated levels of one or more proteins, chitins, elements, amino acids, vitamins, sterols, and / or compounds relative to the same species of insects at the same stage of development fed a diet not containing mushroom biomass.

2. The enriched insects of claim 1, wherein the enriched insects are from the order Coleoptera, Diptera, and / or Orthoptera.

3. The insect enrichment method according to claim 1, wherein the mushroom biomass is derived from Agaricus bisporus (button mushroom). Agaricus bisporus ), Agaricus blazei ( Agaricus blazei ), oyster mushroom ( Pleurotus ostreatus ), Elm Yellow Mushroom ( Pleurotus citrinopileatus Florida Pleurotus ( Pleurotus florida ), lung-shaped lateral ear ( Pleurotus pulmonarius ), King oyster mushroom ( Pleurotus eryngii ), red oyster mushroom ( Pleurotus djamor ), tea tree mushroom ( Agrocybe aegerita ), Elm-shaped pleated umbrella ( Hypsizygus ulmarius ), Chicken leg mushroom ( Coprinus comatus ), Ganoderma lucidum ( Ganoderma lucidum ), Cordyceps militaris ( Cordyceps militaris Cordyceps sinensis Cordyceps sinensis ), Agaricus subrufescens, and Pleurotus ostreatus ( Stropharia rugosoannulata ), Enoki mushrooms Flammulina velutipes ), spotted jade mushroom ( Hypsizygus tessellatus ), True Plum Mushroom ( Hypsizygus marmoreus Poplar mushroom () Cyclocybe aegerita ), Hydrangea ( Sparassis crispa Coral monkey head mushroom () Hericium coralloides ), Hericium erinaceus ( Hericium erinaceus ), Tropical porchia ( Tuber borchii Truffle (Indian truffle) Tuber indicum ), Polyporus umbellatus ( Polyporus umbellatus ), Grifola frondosa ( Grifola frondosa ), sulfur bacteria ( Laetiporus sulphureus ), Ganoderma lucidum ( Ganoderma lucidum ), Nameko mushroom ( Pholiota nameko ), Kuhn mushroom ( Kuehneromyces mutabilis ),straw mushroom( Volvariella volvacea Coral-shaped Hericium erinaceus ( Hericium coralloides ), Hericium erinaceus ( Hericium erinaceus ),mushroom( Lentinula edode s), hairy fungus ( Auricularia polytricha ), Grifola frondosa ( Grifola frondosa ), dark blue light umbrella ( Psilocybe cyanescens ), Cuban psilocybin Psilocybe cubensis ), Psilocybe chinensis ( Psilocybe semilanceata ), Psilocybe nigrum ( Psilocybe mexicana ), Psilocybe nigrum ( Psilocybe muscorum ), Cuban psilocybin Stropharia cubensis ), Yunzhi Cork Fungi ( Trametes versicolor ), Inonotus obliquus ( Inonotus obliquus Yunzhi () Coriolus versicolor ), Enoki mushrooms Flammulina velutipes ), Sanghuang ( Phellinus linteus ).

4. The enriched insect of claim 1, wherein the mushroom biomass comprises mushroom fruiting bodies and / or mycelium and / or stipes.

5. The enriched insects of claim 3, wherein the insects are insect larvae and / or adult insects, wherein the mushroom biomass is derived from Agaricus bisporus (Lange) Imbach. Agaricus bisporus ).

6. The enriched insects of claim 2, wherein the enriched insects are one or more species selected from the group consisting of: Alphitobius diaperinus ( Alphitobius diaperinus ), Armadillidium vulgare ( Alphitobius laevigatus ), Tenebrio molitor ( Tenebrio molitor ), Alphitobius fasciatus ( Tenebrio obscurus ), Diapericus constrictus ( Tenebrio opacus ), Zophobas atratus, Zophobas morio, Hermetia illucens ( Hermetia illucens ), Musca domestica ( Musca domestica ), Acheta domesticus ( Acheta domestica ), Gryllodes sigillatus, Gryllus assimilis, Gryllus bimaculatus ( Gryllus bimaculatus ).​​​​​​​​​ 7. The enriched insect of claim 1, wherein the elements comprise one or more of potassium, calcium, manganese, iron, copper, zinc, or rubidium, the amino acids comprise essential amino acids, the vitamins comprise vitamin Bl, vitamin B2, niacin, and / or nicotinic acid, and the organic compounds comprise melanin or chitin.

8. The nutrient-enriched insect of claim 1, wherein the one or more elements, amino acids, vitamins, compounds, and / or sterols reach a peak maximum value during a finishing period, and the content at the end of the finishing period is lower than the peak maximum value.

9. The enriched insect of claim 1, wherein the one or more elements, amino acids, vitamins, compounds, and / or sterols are selected from the group consisting of 5-hydroxytryptophan, L-tryptophan, serotonin, melatonin, ecdysone, ergosterol, lovastatin, ergothioneine, gamma-aminobutyric acid (GABA), phenolic compounds, cordycepin, erinacin, hericenones, psalliot, dephospho-psalliot, enzymes, B vitamins, macrominerals, trace elements, chitin, beta-glucans, triterpenoids, melanin, tryptophan, phenylalanine, ergothioneine, and ergosterol.

10. The enriched insect of claim 1, wherein the protein content is increased.

11. A method of increasing the levels of elements, amino acids, vitamins, and / or sterols in an insect larva and / or other animal or human, the method comprising: obtaining insect eggs, hatching the eggs into insect larvae, culturing the insects in the presence of a feed comprising mushroom biomass to produce enriched insects, and optionally ingesting the enriched insects directly or indirectly by a human.

12. The method of claim 10, wherein the insect larvae are Coleoptera, Orthoptera, or Diptera insects.

13. The method of claim 10, wherein the mushroom biomass is derived from Agaricus bisporus Agaricus bisporus ).

14. The method of claim 10, wherein the elements comprise one or more of potassium, calcium, manganese, iron, copper, zinc, or rubidium, the amino acids comprise one or more essential amino acids, the vitamins comprise vitamin Bl, vitamin B2, or niacin, and the sterols comprise ergosterol.

15. The method of claim 10, wherein the amino acids comprise one or more of tryptophan, phenylalanine, or ergothioneine.

16. The method according to claim 10, wherein the insects are Coleoptera and / or Diptera and / or Orthoptera insects, which are fed with feed containing mushroom biomass derived from Agaricus bisporus.

17. The method of claim 15, wherein, The elements are one or more of potassium, calcium, manganese, iron, copper, zinc or rubidium, the amino acids are essential amino acids, the vitamins are one or more of vitamin B1, vitamin B2 or niacin, and the sterol is ergosterol.

18. Feed for feeding insects, the feed comprising bran and / or any other cereal and its milled products, pulp of post-cultivation mushroom biomass (waste mushroom substrate) and / or mushroom stems, and feed additives.

19. The feed of claim 18, wherein, The feed comprises about 30-35 wt% wheat bran, 55-60 wt% post-cultivation pulp of mushroom biomass and / or mushroom stems, and 5-15 mass% feed additives, wherein the feed additives comprise brewer's barley spent grain and / or dry feed yeast and / or wet yeast cake, and / or animal protein pulp in the form of slaughterhouse waste, fish waste and / or stale meat food.

20. The feed according to claim 18, wherein the feed comprises about 30-50 wt% wheat bran and / or any other cereal and its milled products and / or brewer's barley spent grain, and further comprises 50-70 wt% of leachate / filtrate of fungal protein crops and / or water after flushing mushrooms and / or water for blanching mushrooms, and / or liquid of in vitro cultivation of mycelium in crops.

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