Insect-derived hydrolysate and production method

Abstract

The present invention relates to a hydrolysate obtained from insects, which contains at least 50% by weight of protein, at least 7% by weight of lipids, and at least 10.5% by weight of ash, these percentages being expressed relative to the dry weight of the hydrolysate, and which has a dry matter content of 15-35% by weight, these percentages being expressed relative to the weight of the hydrolysate. The present invention also relates to a method for preparing the hydrolysate and its use, in particular for improving fish growth.

Description

[Technical Field]

[0001] The present invention relates to hydrolysates obtained from insects, as well as to a process for their preparation and their use in animal feed, in particular fish feed. [Background technology]

[0002] Aquaculture has been a steadily growing business for decades and plays a key role in global food security, but it faces significant challenges due to intensifying production and the importance of using sustainable ingredients with low carbon footprints, particularly when it comes to feed.

[0003] The need to reduce reliance on fishmeal in fish feed production has driven the development of alternative ingredients of marine or terrestrial origin. These new feeds, including functional supplements, can play an important role in the transition to more sustainable practices, thereby reducing environmental impact and improving economic income for fish farmers.

[0004] Therefore, there is a need to find alternatives that can optimize fish development and obtain optimal growth. Summary of the Invention [Problem to be solved by the invention]

[0005] The inventors' research has demonstrated that it is possible to develop insect-derived hydrolysates that can significantly improve fish growth performance even when added in small amounts.

[0006] Furthermore, the inventors have demonstrated that this hydrolysate is also able to improve the consistency of feces, a parameter that is important to consider in recirculating aquaculture systems (RAS) since it has a significant impact on water quality. Indeed, waste must be kept to a minimum and easily removable mechanically to avoid filter degradation. [Means for solving the problem]

[0007] The present invention relates to a hydrolysate obtained from insects, which comprises at least 50% by weight of protein, at least 7% by weight of lipids and at least 10.5% by weight of ash, these percentages being expressed relative to the dry weight of the hydrolysate, and which has a dry matter content of 15 to 35% by weight, these percentages being expressed relative to the weight of the hydrolysate.

[0008] "Hydrolysate" refers to a composition containing hydrolyzed protein. Protein hydrolysates are obtained by enzymatic hydrolysis of proteins. Enzymatic hydrolysis of proteins corresponds to the cleavage of peptide bonds by enzymes at specific cleavage sites, releasing proteins and amino acids of reduced size.

[0009] The hydrolysate according to the invention is in particular a hydrolyzed aqueous composition, i.e. with a humidity level of 65-85%, preferably 70-80%, more preferably 75-80%. Throughout the application, the terms "hydrolyzed aqueous composition" and "hydrolysate" are considered equivalent.

[0010] It should be noted that within the scope of this application, unless otherwise specified, the ranges of values ​​stated are understood to include the boundary values.

[0011] As mentioned above, the hydrolyzed aqueous composition comprises 15-35% by weight dry matter, preferably 20-32% by weight, more preferably 20-25% by weight dry matter, based on the weight of the hydrolyzate.

[0012] It should be noted that within the scope of this application, the terms "weight percentage" and "mass percentage" are synonymous.

[0013] Preferably, the humidity level is measured according to the method of EC Regulation 152 / 2009 as detailed in Example 2.

[0014] Throughout this application, if no date for a regulation, standard, or directive is specified, the regulation, standard, or directive in force on the filing date applies.

[0015] By "insect" is meant insects in any developmental stage, including adults, larvae, nymphs, etc. Preferably, the insects used in the methods of the present invention are edible.

[0016] More specifically, the insect may be selected from the group consisting of Coleoptera, Diptera, Lepidoptera, Isoptera, Orthoptera, Hymenoptera, Blattodea, Hemiptera, Heteroptera, Neuroptera, Ephemeroptera and Macroptera, preferably selected from Coleoptera, Diptera, Orthoptera, Neuroptera and Lepidoptera.

[0017] Preferably, the insect is selected from the group consisting of Tenebrio molitor, Hermetia illucens, Galleria mellonella, Alphitobius diaperinus, Zophobas morio, Blattera fusca, Tribolium castaneum, Rhynchophorus ferrugineus, Musca domestica, Chrysomya megacephala, Locusta migratoria, Schistocerca gregaria, Acheta domesticus, and Samia ricini.

[0018] More preferably, in the hydrolysate according to the invention, the insects are Coleoptera.

[0019] Preferably, the Coleoptera used belongs to the Tenebrionidae, Melolonthidae, Dermestidae, Coccinellidae, Cerambycidae, Carabidae, Buprestidae, Cetoniidae, Curculionidae or mixtures thereof, more preferably the insect belongs to the Tenebrionidae.

[0020] More preferably, they are from the following Coleoptera: Teneworms, Mealworms, Superworms, Teneworms, Red Flour Beetles and Red Palm Weevils, or mixtures thereof, even more preferably Teneworms and Teneworms.

[0021] Preferably, insects are farmed rather than collected from the wild.

[0022] For example, insects are raised on insect farms. Raising insects on dedicated farms not only helps to control and eliminate the risks associated with insect-borne diseases, but also limits the risks associated with the toxicity of insect-derived feed products due to, for example, the presence of pesticides. Furthermore, farming allows for control over the quality of the insect supply and reduces supply costs.

[0023] Advantageously, the hydrolysate according to the invention comprises a fibre content of between 1 and 5% by weight of the dry weight of the hydrolysate.

[0024] Preferably, fiber content is measured according to AOAC method 985.29 as detailed in Example 2.

[0025] Advantageously, the hydrolysate contains between 2 and 5%, preferably between 2.5 and 4.5%, and more preferably between 3 and 4% by weight of fibre relative to the dry weight of the hydrolysate.

[0026] Advantageously, the fibre comprises chitin.

[0027] In this application, "protein" refers to the amount of crude protein. Methods for quantification of crude protein are well known to those skilled in the art. Examples include the Dumas method and the Kjeldahl method. Preferably, the Kjeldahl method is used as detailed in Example 2. Unless otherwise specified, "protein" refers not only to proteins but also to peptides and free amino acids.

[0028] Advantageously, the hydrolysate comprises between 50 and 80% by weight, preferably between 55 and 70% by weight, more preferably between 60 and 65% by weight of protein relative to the dry weight of the hydrolysate.

[0029] Advantageously, the hydrolysate comprises at least 20% by weight of essential amino acids, preferably between 20 and 55% by weight, more preferably between 40 and 50% by weight of essential amino acids, these weight percentages being expressed relative to the total weight of protein in the hydrolysate.

[0030] "Total protein weight" or "protein weight" means the weight of crude protein present in the hydrolysate, unless otherwise indicated as to the nature of the protein, and therefore includes water-soluble and insoluble protein.

[0031] Preferably, the content of essential amino acids is measured according to ISO 13903:2005 and EC Regulation 152 / 2009 (for tryptophan).

[0032] "Essential amino acids" means the following amino acids: histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine. It should be noted that other amino acids may also be essential for some species, for example, arginine for fish.

[0033] Advantageously, the hydrolysate comprises at least 1% by weight of free amino acids, preferably between 2 and 15% by weight, more preferably between 3 and 11% by weight of free amino acids, these percentages being expressed relative to the dry weight of the hydrolysate.

[0034] Preferably, the content of free amino acids is measured according to the ISO 13903:2005 method as detailed in Example 2.

[0035] The five most abundant total amino acids are glutamic acid, aspartic acid, leucine, lysine, and tyrosine.

[0036] These amino acids are particularly beneficial for fish health (e.g., glutamate for gut health and hormone secretion, leucine for muscle protein synthesis).

[0037] The hydrolysate according to the invention preferably comprises 40-60% by weight of insoluble protein, this weight percentage being expressed relative to the total weight of protein, whereas the hydrolysate according to the invention preferably comprises 40-60% by weight of water-soluble protein, this weight percentage being expressed relative to the total weight of protein.

[0038] Advantageously, the hydrolysate according to the invention comprises water-soluble proteins, 75% of which have a size of less than 12400 g / mol, relative to the total weight of the water-soluble proteins.

[0039] "Water-soluble protein" refers to a protein (or crude protein) that dissolves in a solution consisting of 30% acetonitrile, 70% ultrapure water, and 0.1% trifluoroacetic acid ("ACN / water / TFA solution"), where these percentages are by volume relative to the total volume of the solution, as detailed in Example 2.

[0040] By "insoluble protein" is meant a protein that is not soluble in the ACN / water / TFA solution as detailed in Example 2.

[0041] Advantageously, the hydrolysate has a content of more than 80% by weight, preferably between 85 and 98% by weight, more preferably between 90 and 95% by weight, of water-soluble proteins having a size of less than 12400 g / mol relative to the total weight of water-soluble proteins.

[0042] Advantageously, the hydrolysate has a content of more than 40% by weight, preferably between 50 and 70% by weight, more preferably between 55 and 60% by weight, of water-soluble proteins having a size of less than 555 g / mol relative to the total weight of water-soluble proteins.

[0043] Preferably, protein size is measured by HPLC-SEC as detailed in Example 2.

[0044] Due to the small size of the proteins in the hydrolysate according to the invention, the hydrolysate is highly digestible.

[0045] Advantageously, the hydrolysate has a pepsin and / or ileal digestibility of at least 95% by weight of the total weight of protein, preferably at least 96% by weight, more preferably at least 98% by weight, in particular at least 99% by weight.

[0046] Preferably, pepsin digestibility is measured according to AOAC 971.09, AOAC 992.15, AOAC 990.03, and AOCS Ba 4e-93 methods as detailed in Example 2.

[0047] Ileal digestibility is measured according to the BOISEN and Dumas methods as detailed in Example 2.

[0048] As mentioned above, the hydrolysate according to the invention comprises at least 7% by weight of lipids, expressed relative to the dry weight of the hydrolysate. Preferably, the hydrolysate comprises 7 to 20% by weight of lipids, preferably 8 to 18% by weight, more preferably 9 to 16% by weight of lipids, relative to the dry weight of the hydrolysate.

[0049] Methods for determining fat (lipid) content are well known to those skilled in the art. Preferably, the determination of this content is carried out according to the method of EC Regulation 152 / 2009, as detailed in Example 2.

[0050] As mentioned above, the hydrolysate according to the invention comprises an ash content of at least 10.5% by weight, expressed relative to the dry weight of the hydrolysate. Advantageously, the hydrolysate comprises an ash content of 11 to 35%, preferably 12 to 30%, more preferably 15 to 28% by weight, relative to the dry weight of the hydrolysate.

[0051] Preferably, the ash content is measured according to the method of EC Regulation 152 / 2009 as detailed in Example 2.

[0052] Advantageously, the hydrolysate according to the invention comprises one or more additives chosen from preservatives and / or pH-lowering agents.

[0053] The additives added to the hydrolysate are intended to stabilize the hydrolysate, allowing it to be stored at 30°C for periods of more than 6 months, for example 6-12 months.

[0054] Preferably, the preservative is selected from sodium formate, sorbic acid, formic acid, potassium diformate, calcium formate, sodium bisulfite, potassium sorbate, acetic acid, sodium diacetate, calcium acetate and prenyl acetate.

[0055] The preservative is preferably placed in the hydrolysate at a concentration of 0.05 to 3% by weight, preferably 0.1 to 2% by weight of the hydrolysate.

[0056] "pH-lowering agent" means a substance capable of lowering the pH of an aqueous solution.

[0057] Preferably, the pH-lowering agent is selected from organic or inorganic acids, preferably inorganic acids such as phosphoric acid, citric acid, fumaric acid, acetic acid, sorbic acid or propionic acid.

[0058] Advantageously, the pH-lowering agent is introduced in an amount necessary and sufficient to lower the pH to between 2 and 4, preferably between 2.5 and 3.5, for example between 2.8 and 3.2.

[0059] The present invention also relates to a method for preparing a hydrolysate from insects, comprising the following steps: a) Separation of the cuticle and soft parts of the insect; b) separating the soft parts of the insect into an aqueous fraction, an oily fraction and a solid protein fraction; and c) Enzymatic hydrolysis of the solid protein fraction The present invention relates to a method comprising:

[0060] The process according to the invention gives a hydrolysate according to the invention.

[0061] The hydrolysates and insects, including advantageous and preferred aspects thereof, are as described above.

[0062] Therefore, in the method according to the invention, the enzymatic hydrolysis is carried out on the solid protein fraction, and therefore after the soft part has been separated into three fractions. The inventors have demonstrated that carrying out the hydrolysis step on the solid protein fraction after the separation of the soft part into three fractions has various advantages over methods in which the hydrolysis is carried out before the separation into the three fractions, in particular improving the profitability of the process and allowing the other fractions to remain untouched.

[0063] Preferably, the method according to the invention further comprises an insecticidal step of killing insects before the step of separating the cuticle from the soft parts.

[0064] This insecticidal step is described in more detail below in step 1 of the method of the present invention.

[0065] The cuticle is the outer layer (or exoskeleton) secreted by the insect's cuticle. It generally consists of three layers: the epicuticle, the exocuticle, and the endocuticle.

[0066] "Soft parts" refers to the flesh (including, among other things, muscle and internal organs) and body fluids (including, among other things, biofluids, water, and hemolymph) of an insect. In particular, soft parts do not consist of body fluids of an insect.

[0067] Any type of suitable separator may be used to separate the cuticle from the soft parts of the insect, such as a belt separator or a twin screw separator.

[0068] The separation of the cuticle from the soft parts of the insect is described in more detail below in step 2 of the method of the present invention.

[0069] Advantageously, the method according to the invention further comprises a step of maturing the soft parts of the insect between the step of separating the cuticle from the soft parts and the step of separating the soft parts of the insect into an oily fraction, a solid fraction and an aqueous fraction.

[0070] By "process for maturing the soft parts of insects" is meant in particular a process in which the soft parts of insects are heated and / or agitated.

[0071] This step is described in more detail below in step 3 of the method according to the present invention.

[0072] The separation into three fractions aims to recover three fractions from the insect soft parts obtained in step 2 or optionally in step 3: a solid fraction, an aqueous fraction and an oily fraction.

[0073] The separation into these three fractions is described in more detail below in step 4 of the method of the present invention.

[0074] The solid protein fraction (also called the "protein cake") is then subjected to an optional dilution step before being enzymatically hydrolyzed.

[0075] The dilution step aims to reduce the dry matter content preferably to a content of 15-35% by weight.

[0076] The enzymatic hydrolysis step is carried out only on the solid fraction obtained after separation into three fractions. Preferably, at least one proteolytic enzyme is used, preferably a protease, more preferably at least two proteases, which are of different nature. In the present application, the names or suffixes "peptidase" and "protease" are used interchangeably to refer to enzymes that dissolve the peptide bonds of proteins.

[0077] Advantageously, aminopeptidases and serine endopeptidases are used.

[0078] Such an enzymatic hydrolysis step of the solid protein fraction (protein cake) is described in more detail below in step 6 of the process according to the invention. [Brief explanation of the drawings]

[0079] [Figure 1]Figure 1a shows a graph comparing the overall growth performance of rainbow trout fed the control diet (CTRL) for 84 days with that of rainbow trout fed the experimental diet (hydrolysate) of the present invention. The final weights (in grams) of trout obtained from the two diets tested are compared. The diets and parameters measured are described in more detail in Example 3 below. Figure 1b shows a graph comparing the overall growth performance of rainbow trout fed the control diet (CTRL) for 84 days with that of rainbow trout fed the experimental diet (hydrolysate) of the present invention. The final body lengths (in centimeters) of trout obtained from the two diets tested are compared. The diets and parameters measured are described in more detail in Example 3 below. Figure 1c shows a graph comparing the overall growth performance of rainbow trout fed the control diet (CTRL) for 84 days with that of rainbow trout fed the experimental diet (hydrolysate) of the present invention. The weight gains (in percent) of trout measured from the two diets tested are compared. The diets and parameters measured are described in more detail in Example 3 below. Figure 1d shows a graph comparing the overall growth performance of rainbow trout fed the control diet (CTRL) for 84 days with that of rainbow trout fed the experimental diet (hydrolysate) of the present invention. The specific growth rates (units: % / day) of the trout measured using the two diets tested are compared. The diets and parameters measured are described in more detail in Example 3 below. Figure 1e shows a graph comparing the overall growth performance of rainbow trout fed the control diet (CTRL) for 84 days with that of rainbow trout fed the experimental diet (hydrolysate) of the present invention. The daily feed intake (vertical axis: percentage of body weight per day) of the trout measured using the two diets tested is compared. The diets and parameters measured are described in more detail in Example 3 below. Figure 1f shows a graph comparing the overall growth performance of rainbow trout fed a control diet (CTRL) for 84 days with that of rainbow trout fed an experimental diet (hydrolysate) of the present invention. The feed conversion ratios of the trout measured with the two diets tested are compared. The diets and parameters measured are described in more detail in Example 3 below. [Figure 2]FIG. 2 is a bar graph comparing slaughter yields of salmon fed a diet of the present invention containing insect-derived hydrolysate with a control diet. [Figure 3] 3 is a bar graph comparing the fecal consistency of salmon fed a diet of the present invention containing insect-derived hydrolysates with that of salmon fed a control diet, with the vertical axis representing the percentage distribution of feces according to consistency (i.e., a score of 1 or 2). [Figure 4] Figure 4a shows a comparison of the overall growth performance of Atlantic salmon fed the control diet (CTRL2) for 58 days with that of Atlantic salmon fed the experimental diet (hydrolysate) according to the present invention. The final weights (in grams) of the salmon obtained with the two diets tested are compared. The diets and parameters measured are described in more detail in Example 5 below. Figure 4b shows a comparison of the overall growth performance of Atlantic salmon fed the control diet (CTRL2) for 58 days with that of Atlantic salmon fed the experimental diet (hydrolysate) according to the present invention. The final body length (in centimeters) of the salmon obtained with the two diets tested are compared. The diets and parameters measured are described in more detail in Example 5 below. Figure 4c shows a comparison of the overall growth performance of Atlantic salmon fed the control diet (CTRL2) for 58 days with that of Atlantic salmon fed the experimental diet (hydrolysate) according to the present invention. The specific growth rates (in % / day) of the salmon measured using the two diets tested are compared. The diets and parameters measured are described in more detail in Example 5 below. Figure 4d shows a comparison of the overall growth performance of Atlantic salmon fed a control diet (CTRL2) for 58 days with that of Atlantic salmon fed an experimental diet (hydrolysate) according to the present invention. The feed conversion ratios of the salmon measured using the two diets tested are compared. The diets and parameters measured are described in more detail in Example 5 below. DETAILED DESCRIPTION OF THE INVENTION

[0080] The method according to the invention will now be described in more detail.

[0081] Details of the method for treating insects according to the present invention Step 1: Insecticide Killing step 1 can advantageously be carried out by heat shock, such as blanching, which kills the insects while reducing the microbial load (reducing the risk of spoilage and health risks) and inactivating internal enzymes in the insects that can cause autolysis and therefore rapid browning of the insects.

[0082] For blanching, the insects, preferably the larvae, are immersed in boiling water for 2 to 20 minutes, preferably 5 to 15 minutes, preferably at a temperature of 87 to 100°C, preferably 92 to 95°C.

[0083] The amount of water added during blanching is determined as follows: the ratio of the volume of water (ml) to the weight of the insect (g) is preferably 0.3 to 10, more preferably 0.5 to 5, even more preferably 0.7 to 3, and even more preferably about 1.

[0084] For blanching, insects, preferably larvae, are blanched with water or steam (nozzle or steam bed) at temperatures between 80 and 105°C, preferably between 87 and 105°C, more preferably between 95 and 100°C, and even more preferably between 98°C, or with water (by spray nozzle) at temperatures between 90 and 100°C, preferably between 92 and 95°C, or in a mixed mode (water + steam) at temperatures between 80 and 130°C, preferably between 90 and 120°C, more preferably between 95 and 105°C, and even more preferably at 98°C. When blanching insects only with steam, it is advantageous to carry out this in a forced circulation steam heat treatment device ("forced steaming"). The residence time in the blanching chamber is between 5 seconds and 15 minutes, preferably between 1 and 7 minutes.

[0085] Advantageously, after the killing step 1, the insects are used directly to carry out the step 2 of separating the cuticle from the soft parts of the insect, i.e. the insects are not subjected to any treatment such as crushing, freezing or dehydration between steps 1 and 2.

[0086] Step 2: Separation of the cuticle from the soft parts of the insect Step 2 aims to separate the cuticle from the soft parts of the insect.

[0087] The separation of the cuticle from the soft parts of the insect can be carried out using any type of suitable separator.

[0088] According to a first embodiment, the separation of the cuticle and the soft part is carried out using a filter press.

[0089] Advantageously, the filter press used in the insect treatment method according to the invention is a belt filter press.

[0090] A belt filter press has two perforated clamping belts (also called "filter fabrics") between which insects are placed, and the soft parts of the insects are forced through the holes in the clamping belts by pressure, while the hard parts of the insects remain between the two perforated clamping belts.

[0091] Those skilled in the art can determine the diameter of the holes in the fastening belt and the pressure to be applied that will allow separation of the cuticle from the soft parts of the insect.

[0092] According to a second embodiment, the separation of the cuticle and the soft part is carried out using a belt separator.

[0093] For example, the belt separator can include a fastening belt and a perforated drum, with the fastening belt surrounding at least a portion of the perforated drum.

[0094] A clamping belt allows the insects to be transported and pressed against a perforated drum, with the soft parts of the insects being forced through the holes in the drum by pressure, and the hard parts of the insects (cuticles) remaining on the outside of the drum.

[0095] The cuticle can then be collected using a scraper knife.

[0096] With regard to pressure, one skilled in the art can determine the pressure to apply that will allow separation of the cuticle from the soft parts of the insect.

[0097] According to a third embodiment, the separation of the cuticle and soft parts is carried out using a twin-screw separator. This separator has twin screws inside a perforated drum. The soft parts of the insects pass through the holes in the drum, while the cuticle is carried along the two screws to the outside of the separator.

[0098] Advantageously, the diameter of the holes in the drum is between 0.5 and 3 mm, preferably between 1 and 2 mm.

[0099] This insect separation process differs from conventional squeezing in that it allows for the (clean) separation of the soft parts and cuticle of the insect without separating the hemolymph from the solid fraction.

[0100] The soft part obtained in step 2 contains lipids in an amount of 20 to 50% by weight, preferably 30 to 40% by weight, based on the dry weight of the soft part.

[0101] Additionally, the soft portion comprises at least 45%, preferably at least 48%, and more preferably at least 50% by weight of protein based on the dry weight of the soft portion.

[0102] Step 3 (optional): Curing the soft parts of the insects The soft parts of the insects are then optionally subjected to a ripening process in a tank.

[0103] Advantageously, maturation is carried out for a period of between 15 minutes and 3 hours, preferably 1 hour.

[0104] Advantageously, maturation is carried out at a temperature between 65 and 100°C, preferably between 85 and 100°C, more preferably at a temperature of about 90°C.

[0105] This step may also be subjected to stirring.

[0106] This step makes it easier to separate the soft parts of the insect in step 4 below.

[0107] Preferably, the method according to the invention comprises such a step.

[0108] Notably, this process does not require any dilution of the insect soft parts with a solvent such as water.

[0109] Step 4: Separation of the soft part into solid, aqueous and oily fractions This step aims to recover three fractions from the insect soft parts obtained in step 2 or step 3: a solid fraction, an aqueous fraction and an oily fraction.

[0110] According to a first embodiment, this step of separating the soft parts is carried out in two substeps.

[0111] In the first sub-step, the soft parts of the insects are decanted using a two-phase decanter to obtain a solid fraction and a liquid fraction.

[0112] In a second sub-step, the liquid fraction is centrifuged to recover an oily fraction and an aqueous fraction.

[0113] Advantageously, a plate centrifuge is used in this second substep.

[0114] According to a second embodiment of step 4, the soft parts of the insects are decanted using a three-phase decanter to directly obtain the aqueous, oily and solid fractions.

[0115] A suitable three-phase decanter is, for example, the Tricanter® from Flottweg, or a three-phase decanter from GEA, such as the CA225-03-33 decanter.

[0116] Advantageously, the separation of the soft part is carried out according to the second embodiment.

[0117] In fact, the use of a three-phase decanter results in a particularly effective phase separation: more specifically, the resulting solid fraction has a high dry matter content, the aqueous fraction is almost free of insoluble sediments (originating from the solid fraction) and oil, and the oily fraction is almost free of insoluble sediments (originating from the solid fraction) and water.

[0118] Step 5 (optional): Dilution The solid fraction (protein cake) obtained at the end of step 4 is then optionally diluted with water to reduce the dry matter content of this fraction. Specifically, the dilution step is carried out by adding water, such as tap water or soft water. This step is preferably carried out with stirring. Furthermore, heating, for example to 30-80°C, preferably 40-70°C, more preferably 50-70°C, may be carried out.

[0119] At the end of the dilution step, the diluted solid fraction has a dry matter content of 15-35% by weight, preferably 20-25% by weight.

[0120] ·Step 6: Enzyme hydrolysis The purpose of enzymatic hydrolysis is to reduce the size of the proteins in the protein cake, making them more soluble and digestible.

[0121] Enzymatic hydrolysis is carried out by at least one proteolytic enzyme, preferably a protease. In the present application, the names or suffixes "peptidase" and "protease" are used interchangeably to denote enzymes that dissolve the peptide bonds of proteins.

[0122] Advantageously, the hydrolysis is carried out for 1 to 6 hours, preferably 2 to 4 hours, at a temperature of 45 to 75°C, preferably 50 to 65°C, and at a pH of 6 to 8, preferably 7 to 7.5.

[0123] The enzymatic hydrolysis may be carried out with a single protease or with an enzyme mixture comprising at least one protease, more preferably an enzyme mixture comprising multiple proteases.

[0124] Preferably, the protease is selected from the group consisting of aminopeptidases, metallocarboxypeptidases, serine endopeptidases, cysteine ​​endopeptidases, aspartic endopeptidases, and metalloendopeptidases. Advantageously, the enzyme can be chosen from: [Table 1]

[0125] Advantageously, the enzymatic hydrolysis step is carried out using a mixture comprising or consisting of an endopeptidase and an aminopeptidase, more preferably a serine endopeptidase and an aminopeptidase.

[0126] The enzyme or enzyme mixture is dosed in an amount ranging from 2.5 to 31 g / kg of the dry weight of the solid protein fraction, preferably in an amount ranging from 4 to 21 g / kg of the dry weight of the solid protein fraction.

[0127] In terms of enzyme activity, the amount of enzyme or enzyme mixture introduced corresponds to an activity of 40-5000 IU / Kg of dry weight of the solid protein fraction.

[0128] More specifically, the amount of seriendopeptidase added corresponds to an activity of 40 to 60 IU / Kg based on the dry weight of the solid protein fraction.

[0129] Similarly, the amount of aminopeptidase added corresponds to an activity of 2500-5000 IU / Kg of the dry weight of the solid protein fraction.

[0130] Preferably, the process according to the invention further comprises the step of stabilising the solid protein fraction after hydrolysis obtained in step c) by adding one or more additives selected from preservatives and / or pH-lowering agents.

[0131] The additives are as described above, including advantageous and preferred embodiments thereof.

[0132] The present invention also relates to the use of the hydrolysates according to the invention in animal feed.

[0133] Preferably, the animal is selected from livestock, pasture animals, poultry animals, or aquatic animals.

[0134] Advantageously, the domestic animals are selected from dogs, cats, birds, rodents, terrestrial reptiles and fish.

[0135] Advantageously, the grazing animals are selected from cattle, sheep, goats, pigs, horses, camelids and deer.

[0136] Advantageously, the poultry animals are selected from chickens, turkeys, ducks, geese, pigeons, quails, pheasants and ostriches.

[0137] Advantageously, the aquatic animals are selected from seabirds, cetaceans, marine reptiles, crustaceans (such as shrimp), fish, preferably farmed fish.

[0138] Preferred farmed fish include sea bass, sea bream, European sea bream, sturgeon, corvina, sea bass, salmon, tilapia, turbot, and trout.

[0139] Preferably, the farmed fish are selected from the Salmonidae family, which includes salmon and trout.

[0140] Preferably, the fish belongs to the genus Salmo, Salvelinus, Onchorynchus and / or Hucho, more preferably to the genus Salmo.

[0141] Particularly preferred species in the present invention are Salmo salar (Atlantic salmon), Salmo trutta (brown trout or European brown trout), Oncorhynchus kisutch (Pacific salmon), Oncorhynchus tshawytscha (Chinook salmon), Onchorynchus mykiss (rainbow trout), and Salvelinus alpinus (Arctic char).

[0142] Farmed fish are fish raised in tanks or cages in freshwater or saltwater. The environment, water quality, and diet are monitored.

[0143] For example, closed recirculating aquaculture systems (recirculating aquaculture systems) can be used to farm fish.

[0144] Preferably, in the use according to the invention, the feed is in the form of granules, flakes, pate, kibble or treats.

[0145] Advantageously, the feed is in the form of granules or flakes, which are particularly suitable for feeding aquatic animals such as fish.

[0146] In addition to the hydrolysate, such feeds also contain other ingredients suitable for fish feed, such as fish oil or animal meals such as krill meal, fish meal, or squid meal; cereal flours such as wheat flour; oilseed meals such as soybean meal or pea meal; vegetable oils such as rapeseed oil; gelling agents such as guar gum; protein concentrates; wheat or corn gluten; vitamin and mineral premixes; or other ingredients suitable for fish feed.

[0147] Feed in the form of pate or kibble is particularly suitable for feeding domestic animals such as dogs and cats.

[0148] Advantageously, in the use according to the invention, the feed comprises between 1% and 20% by weight of hydrolysate, the percentages by weight being indicated relative to the weight of the feed.

[0149] Advantageously, the feed comprises from 1 to 10% by weight of hydrolysate relative to the weight of the hydrolysate, preferably from 1 to 8% by weight, more preferably from 1 to 5% by weight, the percentages being expressed relative to the weight of the feed.

[0150] Fish feed is most often prepared in the form of granules.

[0151] The production of granules is well known to those skilled in the art and is in particular obtained by extrusion.

[0152] According to a first embodiment, the hydrolysate is mixed with the other ingredients that make up the feed, as mentioned above, before extrusion or molding.

[0153] According to a second embodiment, the other ingredients of the feed are first extruded and then the hydrolysate is introduced in the form of a coating that is applied under vacuum.

[0154] The techniques and conditions for coating granules are well known to those skilled in the art.

[0155] The use according to the invention is advantageously carried out to improve the growth of farmed fish.

[0156] By adding the hydrolysate according to the present invention to fish feed, the weight gain and / or size (body length) of these fish can be significantly improved, preferably improving the weight gain and size of the above fish as shown in Example 3 and FIG. 1.

[0157] More specifically, the use of the hydrolysates of the present invention results in the following in farmed fish, especially salmonids: -Increased weight gain, -Increase in specific growth rate, -Decreased feed conversion ratio, and / or -Improved slaughter yield.

[0158] Preferably, the use of the hydrolysate of the present invention can increase weight gain, increase specific growth rate and decrease feed conversion ratio in farmed fish, especially salmonids.

[0159] The hydrolysates, farmed fish and salmonids according to the invention are more particularly those described above, including advantageous and preferred embodiments thereof.

[0160] The use according to the invention is advantageously carried out to improve the consistency of feces.

[0161] "Fecal consistency" refers to the degree of cohesion of the materials that make up feces. This consistency can be measured on a scale of 1 to 4 as shown in Example 4.

[0162] "Improvement of consistency" means that the cohesiveness of the material that makes up the feces increases, in particular the proportion of solid, hard feces (e.g., score 1) increases and the proportion of liquid feces (diarrhea, loose digesta, score 3-4) decreases.

[0163] High fecal consistency is advantageous for aquaculture applications, which require high water quality to avoid biofilter degradation.

[0164] The present invention also relates to an animal feed containing 1 to 20% by weight of a hydrolysate according to the invention, the weight percentage being given relative to the weight of the feed. Other characteristics and advantages of the present invention will become apparent from the following examples, given by way of example only, and with reference to the following figures: [Example]

[0165] Example 1: Method for preparing a hydrolysate according to the present invention First, 1 kg of T. molitor larvae is steamed and then cut through a twin-screw separator, allowing for the separation of the cuticle and soft parts (flesh).

[0166] The resulting soft portion is placed in an aging tank and aged for 1 hour at 90°C while stirring. The heated soft portion is then separated through a tricanter (three-phase decanter) to obtain an oily fraction, an aqueous fraction, and a solid protein fraction corresponding to the protein cake.

[0167] Therefore, the protein cake is diluted with water to reduce the dry matter from 42% to 22%.

[0168] A hydrolysis step is then carried out at 60°C for 2 hours using the following enzymes: -Alcalase: administered at a concentration of 40-60 IU / kg of dry weight of protein cake; -Flavorzyme: Add at a concentration of 2500-5000 IU / kg of dry weight of protein cake.

[0169] The enzymes are then inactivated at 90°C for 30 minutes, and the resulting hydrolysate is cooled to 40°C before adding additives to stabilize it and obtain a pH below 2.9. The additives used may include preservatives and / or pH-lowering agents.

[0170] Preservatives may be selected from sodium formate, sorbic acid, formic acid, potassium diformate, calcium formate, sodium bisulfite, potassium sorbate, acetic acid, sodium diacetate, calcium acetate and prenyl acetate.

[0171] The pH-lowering agent may be selected from organic or inorganic acids, preferably inorganic acids such as phosphoric acid, citric acid, fumaric acid, acetic acid, sorbic acid or propionic acid.

[0172] Finally, a filtration step is performed to remove any remaining cuticular particles.

[0173] The resulting finished product is a hydrolysate (or hydrolyzed aqueous composition) with a dry matter content of 20-25%.

[0174] Example 2: Characterization of the hydrolysates according to the invention The properties of the hydrolysate obtained in Example 1 are as follows:

[0175] 1.Analysis 1.1 Measuring humidity levels The humidity level was measured according to the method of EC Regulation 152 / 09.

[0176] 1.2 Protein measurement Protein content was determined according to the Kjeldahl method of EC Regulation 152 / 2009 with an N to protein conversion factor of 6.25.

[0177] 1.3 Measurement of lipid content The amount of lipids was determined according to EC Regulation 152 / 2009.

[0178] 1.4 Ash content measurement Ash content was determined according to EC Regulation 152 / 2009.

[0179] 1.5 Measuring the amount of fiber Fiber content (soluble and insoluble) was determined according to AOAC 985.29 method.

[0180] 1.6 Determination of Amino Acid Amounts The amounts of total and free amino acids were determined according to ISO 13903:2005 (for total and free amino acids other than tryptophan) and EC Regulation 152 / 2009 (for tryptophan).

[0181] The amount of free amino acids was determined according to ISO13903:2005.

[0182] 1.7 Pepsin digestibility measurement Pepsin digestibility was measured according to AOAC 971.09 (pepsin digestibility at 0.02% (gravimetric)) and AOAC 992.15, AOAC 990.03, and AOCS Ba 4e-93 (protein combustion) methods.

[0183] 1.8 Measurement of ileal digestibility Ileal digestibility was measured according to the Boysen method (enzyme dosage) and Dumas method (protein content).

[0184] 1.9 Determination of soluble and insoluble protein content The amount of soluble protein was measured by dissolving the above proteins in a solution consisting of 30% acetonitrile, 70% ultrapure water and 0.1% trifluoroacetic acid ("ACN / water / TFA solution"), where these percentages are by volume relative to the total volume of the solution constituting the mobile phase before quantification by HPLC-SEC (a size exclusion chromatography method known to those skilled in the art).

[0185] The amount of insoluble protein was determined by the dry residue obtained after dissolving a sample of the dried hydrolysate in an ACN / water / TFA solution and added to the initial dry weight.

[0186] 1.10 Protein size measurement The size of the protein was determined by HPLC-SEC.

[0187] 1.11 Measurement of mineral content The content of each mineral listed in Table 8 below was measured by ICP / AES (inductively coupled plasma atomic emission spectroscopy).

[0188] 1.12 Vitamin Measurement Thiamine hydrochloride (vitamin B1-HCl) content was determined according to BS EN 14122-2014 method.

[0189] Vitamin B12 (cyanocobalamin) content was determined according to AOAC 952.20 method.

[0190] Riboflavin (vitamin B2) content was determined according to the EN14152:2014 method.

[0191] Niacin (vitamin B3) content was determined according to the EN15652:2009 method.

[0192] Pantothenic acid (vitamin B5) content was determined according to the AOAC 2012.16 method.

[0193] Pyridoxine (vitamin B6) content was determined according to the EN14164:2014 method.

[0194] Phylloquinone (vitamin K1) content was determined according to the EN14148:2003 method.

[0195] 2.Results The properties of the hydrolysates according to the present invention are shown in Tables 2 to 8 below. [Table 2] [Table 3] [Table 4] [Table 5] [Table 6] [Table 7] [Table 8]

[0196] Example 3: Addition of hydrolysate according to the invention to fish feed (rainbow trout) Materials and Methods A fish meal-based diet (CTRL) was formulated with ingredients that meet the known nutritional needs of rainbow trout. Based on this diet, another diet containing the hydrolysate obtained according to Example 1 was prepared (see Table 9 below). [Table 9]

[0197] The two feeds were produced by extrusion according to standard techniques known to those skilled in the art to obtain granules of 2-4 mm size.

[0198] During the extrusion process, the feed is dried and then cooled.

[0199] In the case of granules from CTRL feed, the granules are obtained by extruding a composition prepared by mixing LT fish meal, wheat gluten, soybean concentrate, soybean lecithin, wheat flour, premix, guar gum, and hemoglobin in the amounts shown in Table 8. Then, at the end of extrusion, the extruded granules are cooled and then vacuum coated to add an additional 7% by weight of fish oil.

[0200] In the case of the granules according to the present invention, the granules are obtained by extruding a composition prepared by mixing LT fish meal, wheat gluten, soybean concentrate, soybean lecithin, wheat flour, premix, guar gum, and hemoglobin in the amounts shown in Table 8. Then, at the end of the extrusion, the extruded granules are cooled and then coated twice in succession as follows: - 1st step: The hydrolysate (4% by weight of a composition with a dry matter content of 25%, ie 1% dry matter content relative to the total weight of the granule composition) is added to the extruded granules by vacuum coating. - Second step: The hydrolysate coated granules are dried at low temperature. - Third step: Add 7% by weight of fish oil by vacuum coating.

[0201] b. Growth performance test Each diet was tested in four samples (four tanks per diet).

[0202] The rainbow trout were allowed to acclimate for 15 days before the start of the growth test, after which 25 rainbow trout with an average weight of 20-30 g were randomly distributed to each tank (20 tanks in total). The tanks used were cylindrical glass fiber tanks (volume: 500 L) connected to a recirculating aquaculture system (RAS).

[0203] The tanks were supplied with freshwater at a temperature of 15 ± 1°C with a dissolved oxygen concentration of >7 ± 1 mg / L. Ammonium and nitrite concentrations in the water were monitored daily to ensure they remained below toxic levels. Fish were maintained on a 12-h light / dark cycle.

[0204] Fish were hand-fed once daily (between 8 and 9 am) up to 3% of their daily feed biomass intake until apparent satiety was reached, and this continued for 84 days.

[0205] After 84 days of experimental feeding, all fish were collected from each tank, anesthetized with MS222, and evaluated for growth performance. Parameters measured for growth assessment: PCI(g): Initial weight PCF(g): Final weight TCS (% / day): Specific growth rate = [(ln PCF-ln PCI) / day] × 100 LTI(cm):Initial total length LTF(cm): Final total length GP (%): Weight gain = [(PCF-PCI) / PCI] x 100 RCA: Feed conversion ratio = [total feed intake (g) / GP (g)] PAJ (% of body weight per day): Daily feed intake = (total feed intake / (PCI+PCF) / 2 / day) x 100 [Table 10]

[0206] Values ​​are expressed as mean ± standard deviation (n=4).

[0207] Asterisks (*) indicate significant differences between the experimental and CTRL control groups (Student's t-test, P<0.05, n=4).

[0208] Conclusion: After 84 days of experimental feeding, it was observed that the weight and length of fish fed with granules according to the invention increased significantly compared to fish fed with CTRL granules, when the daily intake was comparable (see Figure 1).

[0209] Thus, the inventors have demonstrated that the use of very small amounts of the hydrolysate according to the invention (1% dry matter) has a significant effect on fish growth.

[0210] Example 4: Addition of hydrolysate according to the invention to fish feed (salmon) Materials and Methods The hydrolysates described in Examples 1 and 2 are used.

[0211] Commercially available granular feed with a size of 2 mm manufactured by Skretting AS (RCX) was used as the basis for producing the two diets described below. These granules contain, among other things, 49% by weight of protein, 22% by weight of lipids, and 9% by weight of ash. Based on this formulation, 1% of the hydrolysate was added by coating to produce the test diets.

[0212] Additionally, in the preparation of these two diets, the granules were coated with water to balance the average moisture content to approximately 13% of the total granule weight.

[0213] A variety of salmon species were tested. A total of 320 Atlantic salmon, each weighing an average of 17 g, were randomly distributed among six 80 L tanks (average weight per tank: 16.63–16.70 g). Three overfeeding tanks were fed each diet.

[0214] The experiment was carried out over a period of three weeks.

[0215] Growth Assessment All fish were weighed individually at the start of the experiment and at harvest (end of the experiment).

[0216] Poop rating At the end of the experiment, 10 fish were randomly selected per tank based on the appearance of their feces.

[0217] Feces consistency was analyzed visually and rated according to a score of 1 to 4: score 1: solid and hard; score 2: semi-solid and soft digesta; score 3: diarrheal and watery digesta; score 4: yellow or non-white clumps of non-feed material / no digesta.

[0218] Digestate corresponds to the residue produced by methanation (anaerobic digestion).

[0219] b.Results Growth Assessment During the experimental period, body weight increased 2.2-fold from 17 g to 38.5-39.5 g, corresponding to a growth rate (SGR) of 3.48-3.58. As can be seen from Figure 2, slaughter yield was significantly lower in the control (82.7%) compared to the hydrolysate-containing diet (83.7%).

[0220] Poop rating The softest consistency was observed in the CTRL group (Table 11, Figure 3, left bar). [Table 11]

[0221] As can be seen from Figure 3 and Table 11, the addition of hydrolysates to salmon feed significantly improved fecal consistency compared to the control diet. In fact, compared to the control diet, the percentage of solid feces (score 1) improved to 70%–80% in the diets containing T. molitor hydrolysates, respectively, compared to 40% in the control diet.

[0222] Due to the low incidence of semi-solid feces, the hydrolysate according to the present invention is useful as an animal feed supplement, especially in aquaculture applications where high water quality is required to avoid degradation of biofilters.

[0223] The experimental data generated in this example confirms that the addition of the hydrolysate according to the present invention to animal feed significantly improves the consistency of feces.

[0224] Example 5: Addition of hydrolysate according to the invention to fish feed (salmon) Materials and Methods The method described in Example 1 was reproduced, resulting in a hydrolysate with the following characteristics: [Table 12]

[0225] A control diet (CTRL2) was formulated with ingredients that meet the known nutritional needs of salmon, and this diet was used as a base to prepare additional diets containing the hydrolysates listed in Table 12 (see Table 13 below). [Table 13] [Table 14]

[0226] The feeds shown in Table 12 were manufactured by extrusion according to standard techniques known to those skilled in the art to obtain granules of size 2 mm.

[0227] The granules were first obtained by mixing soy protein concentrate, fish meal, wheat gluten, potato starch, pea mix, lysine, methionine, monocalcium phosphate, yttrium oxide, and choline chloride in a Spiry 25 dough mixer according to the contents shown in Table 13. Secondly, fish oil and hydrolysate (for feedstuffs according to the invention) were added to the mixture. Thirdly, gelatin activated at 60°C was added to the mixture as well. The paste thus obtained was processed into granules in the cold state using an extruder. The granules were dried at 45°C until the dry matter content reached approximately 93%.

[0228] Thus, according to this embodiment, the hydrolysate is mixed with the other ingredients that make up the feed (granules) before extrusion.

[0229] Each diet was tested in triplicate (three tanks per diet). 180 Atlantic salmon with an average weight of 20.3 ± 0.3 g were randomly distributed among six 80 L tanks. The tanks were connected to a recirculating aquaculture system (RAS). Freshwater was supplied to the tanks at temperatures ranging from 13.3 to 15.3°C (average temperature 14.1°C).

[0230] Each diet was offered at the time of appetite, and the leftovers were collected to accurately calculate the feed conversion ratio (RCA or Feed Conversion Rage).

[0231] Ten salmon were selected per tank for growth performance evaluation, and after 58 days of experimental feeding, these 10 salmon were euthanized by an overdose of anesthetic.

[0232] b.Results Growth Assessment The weight of the 10 selected salmon was measured at the start of the experiment and at the time of harvest (end of the experiment), at which time their body length was measured. [Table 15]

[0233] Values ​​are expressed as mean ± standard deviation (n=3), where standard deviation is equal to standard error.

[0234] Asterisks (*) indicate significant differences between the experimental and CTRL2 control groups (Student's t-test, P<0.05, n=3).

[0235] The above results confirm that the weight of salmon fed the hydrolysate-containing granules increased by 5-fold. Furthermore, when daily feed intake was equalized (RCA of 0.67 for both the CTRL2 group and the hydrolysate-containing group), it was observed that the weight and length of salmon fed the granules of the present invention increased significantly compared to salmon fed the CLTRL2 granules. In fact, a significant difference of 14.3% was obtained in body weight (see Table 15 and Figure 4a).

[0236] Perivisceral fat score and evisceration weight At the end of the experiment, the amount of perivisceral fat in the tested salmon was measured by applying the following visual measurement method. Visceral fat mass was assessed on a scale of 1 to 5 according to the visibility of the pyloric caeca (Fig. 5).

[0237] - Score 1: The pyloric caeca is clearly visible - Score 2: The pyloric caeca is visible - Score 3: The pyloric caeca is visible as a cleft in the visceral fat - Score 4: The pyloric caeca is visible through the visceral fat - Score 5: The pyloric caeca is not visible

[0238] Therefore, the lower the score, the less perivisceral fat there is.

[0239] Additionally, the weight of the gutted salmon was measured.

[0240] The results are shown in Table 16 below. [Table 16]

[0241] These results confirmed that although there was no significant difference between salmon fed with granules from the CTRL2 group and salmon fed with granules according to the present invention, the latter tended to have a higher body weight after gutting.

[0242] Furthermore, no significant differences were observed regarding perivisceral fat deposition.

[0243] Therefore, from all these results it can be concluded that salmon fed with granules according to the invention show significantly greater growth in terms of body weight and length without becoming fat. Therefore, fish feed containing the hydrolysate according to the invention promotes an increase in body length as well as muscle mass in the body without causing fat accumulation around the internal organs. As a result, the hydrolysate according to the invention makes it possible to significantly promote the growth of fish while maintaining their good body condition, thus improving the nutritional value of fish fed with granules containing the hydrolysate according to the invention.

Claims

1. 1. A hydrolysate obtained from insects, comprising at least 50% by weight of protein, at least 7% by weight of lipids, at least 10.5% by weight of ash, these percentages being expressed relative to the dry weight of the hydrolysate, and having a dry matter content of 15-35% by weight, said percentages being expressed relative to the weight of the hydrolysate.

2. The hydrolysate of claim 1 , wherein the insect is a beetle.

3. 10. The hydrolysate of any one of the preceding claims, comprising a fibre content of 1 to 5% by weight of the dry weight of the hydrolysate.

4. 10. The hydrolysate according to any one of the preceding claims, comprising water-soluble proteins, wherein 75% of the water-soluble proteins relative to the total weight of the water-soluble proteins have a size of less than 12,400 g / mol.

5. 10. The hydrolysate according to any one of the preceding claims, comprising one or more additives selected from preservatives and / or pH-lowering agents.

6. 10. A hydrolysate according to any one of the preceding claims, comprising 40 to 60% by weight of insoluble protein, said weight percentages being expressed relative to the total weight of protein.

7. 1. A method for preparing a hydrolysate from insects, comprising the steps of: a) Separation of the cuticle and soft parts of the insect; b) separating the soft parts of the insect into an aqueous fraction, an oily fraction and a solid protein fraction; and c) Enzymatic hydrolysis of the solid protein fraction A method comprising:

8. 8. The method of claim 7, further comprising the step of killing insects before the step of separating the cuticle from the soft parts.

9. 9. The method of claim 7 or 8, further comprising a step of ripening the soft parts of the insect between the step of separating the cuticle from the soft parts and the step of separating the soft parts of the insect into an oily fraction, a solid fraction and an aqueous fraction.

10. 10. The method according to any one of claims 7 to 9, further comprising the step of stabilising the solid protein fraction after hydrolysis obtained in step c) by adding one or more additives selected from preservatives and / or pH-lowering agents.

11. Use of the hydrolysate according to claims 1 to 6 in animal feed.

12. 12. The use according to claim 11, wherein the feed is in the form of granules, flakes, pate, kibble or treats.

13. The use according to claim 11 or 12, wherein the feed comprises from 1% to 20% by weight of hydrolysate, said weight percentages being expressed relative to the weight of the feed.

14. 14. Use according to any one of claims 11 to 13 for improving the growth of farmed fish.

15. 14. Use according to any one of claims 11 to 13 for improving fecal consistency.

16. An animal feed comprising 1 to 20% by weight of a hydrolysate according to claims 1 to 6, said weight percentages being expressed relative to the weight of the feed.

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