Method for producing chitosan from fungal mycelium

Abstract

The invention relates to a method for producing fungal chitosan from a mycelium, which method comprises supplying a reactor with the mycelium and an alkaline material; deacetylating the mycelium; washing the obtained kneaded fungal chitosan; a solubilisation step; optionally filtering the liquid phase; precipitating the liquid phase to obtain a precipitated fungal chitosan and a step of collecting the precipitated fungal chitosan; the method comprising, after the solubilisation step, a step of directionally controlling the liquid phase using a branch channel, during which step the liquid phase is prepared for the filtration step or the precipitation step.

Description

[0001] “Process for producing chitosan from fungal mycelium”

[0002] The present invention relates to a method for producing chitosan from a fungal mycelium, said chitosan being advantageously usable in pharmaceutical, agricultural, food, cosmetic, industrial, oenological and / or environmental applications. The production of chitosan is often carried out from crustacean exoskeletons which contain a significant amount of chitin. Chitin is one of the main elements constituting the exoskeleton of a crustacean and which gives it its rigidity.

[0003] In today's society, it is important to be able to adapt to new trends, such as those in food. Therefore, it is important to be able to produce chitosan from a non-animal source, such as fungal mycelium.

[0004] Mycelium is the vegetative apparatus of fungi or certain filamentous bacteria. According to the present invention, the term mycelium refers to fungal mycelium. It is composed of a set of filaments. Mycelium is an important source of chitin, which is a major component of the cell wall of fungi.

[0005] Mycelium is an alternative to using crustacean exoskeleton because the quality and quantity of chitin present in its walls does not depend on the seasons and the quantity of impurities is lower.

[0006] For example, the mycelium of a fungus such as Aspergillus niger can be used. Aspergillus niger is known primarily as a source of chitin for producing chitosan. The mycelium of Aspergillus niger is a by-product of A. niger fermentation.

[0007] Chitin is a polysaccharide with a structural role due to its strength and flexibility. It is produced by the polymerization of N-acetylglucosamine linked together by a [3-1,4]-type osidic bond. Chitin is widely distributed in both the animal and plant kingdoms. For example, it is present in the exoskeletons of crustaceans, in the walls of fungi, etc. Chitin is a non-toxic and biodegradable insoluble polysaccharide which, when deacetylated, produces chitosan.

[0008] Chitosan is obtained by removing sufficient acetyl groups from chitin. It is a polysaccharide composed of [3-1,4]-linked D-glucosamine and N-acetyl-D-glucosamine whose characteristics vary with the origin of the source, whether animal or plant, and the characteristics also vary within the same source. The quality and quantity of chitosan that can be extracted from the same source, such as mycelium, may vary depending on the batch of mycelium used.

[0009] An increasingly pure and controlled chitosan is sought. Chitosan can be produced directly from chitin. Chitin can be pre-extracted from mycelium. Another possibility is the production of chitosan from mycelium, which is an important source of chitin, as explained above.

[0010] Chitosan is sought after for its multiple applications, such as in cosmetics, dietetics, food processing, etc.

[0011] For example, when chitosan is used in the field of oenology, it must comply with the International Oenological Codex and its constraints. For example, chitosan must be 95% pure, with a glucan content greater than 2% but less than or equal to 5%. Such chitosan has an antibacterial effect and prevents ferric casses and therefore the precipitation of proteins with iron.

[0012] In the agri-food sector, chitosan must meet the criteria of a Novel Food on the European market or the FDA criteria on the American market.

[0013] When chitosan is used in the medical field, in applications such as immune system modulation, glucans play an important role and therefore their percentage should be higher in chitosan.

[0014] When chitosan is used in the dietetic field, a lower percentage of glucans is sought because it is the chitosan that captures the fats.

[0015] Known from the state of the art is document EPI 483299 which describes a process for producing chitin from fungal biomass by an enzymatic process using a [3-1,3-glucanase to obtain chitin. The chitin obtained is then treated by an enzymatic process using a chitin deacetylase to obtain chitosan.

[0016] Unfortunately, this process, using enzymes, is expensive to implement and has limited robustness, which makes its industrial use limited.

[0017] There is therefore a need to develop a process for producing chitosan which allows industrial production at lower costs than the costs generated by a process as described in EPI 483299 and which is more robust. More particularly, the invention provides a process for producing fungal chitosan from a mycelium comprising the steps in order: feeding a conical reactor with the mycelium and an alkaline material; at least partial deacetylation of the mycelium in the conical reactor to obtain a kneaded fungal chitosan; at least one washing of the kneaded fungal chitosan in a washing tank and solubilization of the kneaded fungal chitosan in at least one solubilization tank to obtain a liquid phase containing the solubilized fungal chitosan.

[0018] Such a method for producing chitosan is described in document WO201 3053838. This document describes a non-enzymatic method which involves the production of chitosan on the one hand and glucans on the other hand.

[0019] According to this document, the optimal conditions for chitosan production are not described. The process recommends working under conditions that maximize the amount of glucans produced and therefore recommends using 0.5 to 1.5, or even up to 5 parts by weight of NaOH (dry alkaline matter) per part by weight of mycelium for the deacetylation of the mycelium. While this process is very suitable for glucan extraction, it requires, precisely because of the glucan extraction, working under conditions that do not allow for economically and environmentally efficient chitosan extraction.

[0020] The present invention proposes to overcome the drawbacks of the state of the art. In particular, it intends to provide a robust, reproducible and predictable process allowing industrial exploitation for the production of chitosan from a mycelium, as well as the chitosan thus obtained, that is to say a chitosan whose final characteristics will be predictable and controllable, depending on the applications envisaged.

[0021] Furthermore, the present invention also intends to provide a method which offers great flexibility, great precision and a great degree of freedom for the control on the same production line of the final characteristics of the chitosan product, in particular its glucan content.

[0022] Thus, according to the invention, there is provided a method for producing fungal chitosan from a mycelium comprising the following steps in order: (i) feeding a conical reactor with the mycelium and an alkaline material; (ii) at least partial deacetylation of the mycelium in the conical reactor to obtain a kneaded fungal chitosan; (iii) at least one washing of the kneaded fungal chitosan in at least one washing tank; (iv) solubilization of the kneaded fungal chitosan in at least one solubilization tank to obtain a liquid phase containing the solubilized fungal chitosan; (v) optionally filtration of said liquid phase;(vi) precipitation of said liquid phase in at least one precipitation tank with obtaining a precipitated fungal chitosan and (vii) collection of said precipitated fungal chitosan, said method further comprising, after the solubilization step, a step of directional control of the liquid phase using diversion means, during which the liquid phase is directed towards filtration and / or towards precipitation.;

[0023] By the term "diverting means" according to the present invention, is meant for example multi-way diverting valves, preferably diverting valves with at least three ways. These three-way diverting valves make it possible, on the one hand, to direct the liquid towards a first direction and / or towards a second direction different from the first direction and / or, moreover, to modulate the quantity of liquid directed towards each direction. The choice between the first direction and the second direction is made by the user. These diverting means do not cover a simple valve for opening and / or closing a single liquid flow path.

[0024] By the term "directional control step" according to the present invention is meant a step of choosing the direction of the liquid phase at least between a first direction and a second direction or the choice of the quantity of liquid directed towards each direction. This choice, imperative according to the invention, depends on the desired characteristics of the finished product and the characteristics of the raw materials (e.g. mycelium). The first direction leads to the filtration step, while the second direction leads directly to the precipitation step (without going through the filtration step, which is therefore optional according to the present invention).

[0025] Other characteristics, details and advantages of the invention will emerge from the description and figures given below, without limitation. In particular:

[0026] Figure 1 shows a diagram illustrating the steps of the method according to the invention.

[0027] As illustrated in Figure 1, the method according to the present invention comprises a step of feeding a conical reactor with the mycelium and an alkaline material, which allows the mycelium to come into contact with the alkaline material.

[0028] Advantageously, the mycelium comes from Aspergillus niger.

[0029] Preferably, the alkaline material is in the form of an alkaline solution, in particular in the form of a concentrated alkaline solution. Preferably, the alkaline solution is chosen from an aqueous solution of sodium or potassium hydroxide or their mixture, but is not limited thereto. The alkaline solution may also be a solution of another base compatible with the food industry.

[0030] Preferably, the alkaline solution according to the present invention has a concentration of between 40% and 60%, preferably between 45% and 55%, even more preferably 50%.

[0031] The alkaline solution has a concentration greater than 40% (mass / volume) of alkaline matter providing alkali ions relative to the total mass of the alkaline solution. For example, a sodium hydroxide solution with a concentration of 50% (mass / volume) can be used.

[0032] Advantageously, the alkaline solution according to the invention is preferably an aqueous solution of sodium hydroxide.

[0033] For example, a 50% (mass / volume) sodium hydroxide solution is used, which means that the solution contains 50% NaOH and 50% water to form 100% NaOH solution.

[0034] As illustrated in Figure 1, the method according to the present invention comprises at least partial deacetylation of the mycelium in the conical reactor with obtaining a kneaded fungal chitosan.

[0035] The step of at least partial deacetylation of the mycelium according to the process according to the invention allows a reaction of the mycelium with a reduced quantity of alkaline material compared to the existing process, but also in a more restricted volume of material.

[0036] The reactor in which the mycelium and alkaline matter are fed is for example a 4m conical reactor 3 in which deacetylation occurs. It can advantageously be equipped with a double jacket allowing the reaction medium to be heated up to 120°C. The agitation of the mixture can be ensured by a worm screw mounted on an orbital arm which travels around the periphery of the reactor at a rate of approximately 2 revolutions / min.

[0037] According to one embodiment of the present invention, the deacetylation of the mycelium is carried out at a temperature below 120°C. For example, the temperature increase is gradual.

[0038] In a preferred embodiment, the at least partial deacetylation of a fungal mycelium according to the present invention is carried out at a temperature between 100 and 110°C, preferably between 103 and 107°C, preferentially at a temperature of approximately 105°C.

[0039] In an advantageous embodiment, the at least partial deacetylation of a fungal mycelium according to the present invention is carried out for a period of between 4 and 10 hours, preferably for approximately 6 hours.

[0040] According to one embodiment, the addition of the alkaline material is carried out until a pH adjusted to a value greater than 9.0, preferably greater than 9.5, and more preferably adjusted to 10 is obtained.

[0041] In this way, the reactor is capable of homogenizing the mixture of alkaline material and mycelium but also of raising the temperature of the mixture so as to allow the hydrolysis of the mycelium, the deacetylation of the chitin from the mycelium into chitosan in a restricted reaction volume. The mixture thus obtained contains the kneaded fungal chitosan having a degree of acetylation, that is to say a molar proportion of N-acetyl-D-glucosamine units along the chitosan chains, of 0 to 25%, preferably between 0 and 20%, or even less than 15%. According to the present invention, the deacetylation is carried out directly in the conical reactor, which makes it possible to reduce the energy input necessary for heating the mixture as well as the size of the equipment, while resulting in an advantageous chitosan extraction yield.

[0042] Yield means: the mass ratio expressed as a % of the dry mass of the fraction comprising chitosan (chitosan and its impurities) to the dry mass of the starting mycelium.

[0043] In some embodiments of the invention, the alkaline solution forming essentially the filtrate of the mixture containing the kneaded fungal chitosan is recovered, concentrated, recycled and reused for the deacetylation of the mycelium during another chitosan production. The kneaded fungal chitosan is then transferred, optionally after dilution to facilitate its transfer to the washing tank. The kneaded chitosan can also be filtered to remove a series of contaminants in the liquid phase formed essentially of the alkaline solution.

[0044] As illustrated in Figure 1, the method according to the present invention further comprises (at least one washing of the kneaded fungal chitosan in at least one washing tank.

[0045] The kneaded fungal chitosan is preferably subjected to at least two washes, preferably at least three washes, preferably at least four washes, preferably at least five washes, preferably at least six washes, preferably at least seven washes, in at least one washing tank.

[0046] Advantageously, the at least one wash is a water wash. Advantageously, the step of at least one wash makes it possible to eliminate the [3-glucan.

[0047] Optionally, the method of the invention comprises a dilution and / or filtration step between the step of at least one washing and the solubilization step according to the invention. According to the invention, after the washing step, the method comprises solubilization of the kneaded fungal chitosan in at least one solubilization tank with the production of a liquid phase containing the solubilized fungal chitosan.

[0048] As illustrated in Figure 1, the method according to the present invention further comprises solubilizing the kneaded fungal chitosan in at least one solubilizing tank with the production of a liquid phase containing the solubilized fungal chitosan. Advantageously, the solubilization of the chitosan in the method according to the present invention is carried out by adding an acid, preferably in the form of an acid solution, until a pH of less than 6, preferably less than 5.5 and even more preferably less than 4.5 is obtained. Advantageously, an acid solution having a concentration of between 0.1 and 1 N can be used.

[0049] Solubilization can, for example, be carried out at room temperature or any other temperature that promotes the solubilization of chitosan.

[0050] Advantageously, the solubilization of the fungal chitosan is carried out for a period of between 7 and 9 hours, preferably for approximately 8 hours. In the method according to the present invention, the acid is, for example, in the form of an acid solution, chosen from the group comprising hydrochloric acid, lactic acid, formic acid, glutamic acid, phthalic acid, succinic acid, glycolic acid, citric acid, acetic acid and mixtures thereof, and is preferably acetic acid.

[0051] At the end of solubilization, the mixture obtained is a liquid phase containing the solubilized fungal chitosan and a solid phase.

[0052] As illustrated in Figure 1, the method according to the present invention further comprises, after the solubilization step and preferably directly after the solubilization step, a step of directional control of the liquid phase using diversion means during which the liquid phase is directed towards filtration and / or towards precipitation.

[0053] Preferably, the bypass means is a three-way valve. The first way of the valve directs the liquid to the filtration stage while the second way directs the liquid to the precipitation stage.

[0054] Preferably, the liquid phase is directed to filtration and precipitation in the liquid phase directional control step using the three-way valve. A portion of the liquid phase is directed to the filtration step and another portion of the liquid phase is directed to the precipitation step. The volume of each portion is determined, in particular according to the desired glucan level. The amount of liquid phase containing solubilized fungal chitosan that is directed to the precipitation step allows the glucan level to be varied, with great precision and over a wide range of glucan levels.

[0055] Preferably, when the liquid phase is directed towards filtration and towards precipitation during the directional control step, at least 0.1% by volume of the liquid phase, preferably at least 0.5% by volume of the liquid phase, preferably at least 1% by volume of the liquid phase, preferably at least 1.5% by volume of the liquid phase, preferably at least 2% by volume of the liquid phase, preferably at least 3% by volume of the liquid phase, preferably at least 4% by volume of the liquid phase, preferably at least 5% by volume of the liquid phase, is directed towards precipitation (provided that according to the invention, the remainder of the liquid phase is therefore directed towards filtration).

[0056] In another embodiment according to the present invention, the liquid phase is directed to the filtration step (as illustrated in Fig.l), preferably the total volume of the liquid phase is directed to the filtration step. The direction is given by the three-way valve, the first way for bringing the liquid phase to the filtration step.

[0057] Filtration can be followed by a turbidity measurement to assess the quality of filtration. Turbidity measurement is a measure of the cloudiness of liquids, recognized as a simple and basic indicator of liquid quality.

[0058] In another embodiment according to the present invention, the liquid phase is directed to the precipitation step (as illustrated in Fig.l), preferably the total volume of the liquid phase is directed to the precipitation step. The direction is given by the three-way valve, the second way allowing the liquid phase to be brought to the precipitation step.

[0059] This directional control step advantageously allows the glucan level present in the final chitosan collected to be modulated with great precision and over a wide range. Another advantage of this directional control step is that it provides greater flexibility and a greater degree of freedom during production. Indeed, it is possible to carry out the “non-derived” product synthesis and the “derived” product synthesis on the same production line according to the method of the invention, thus saving space in the production premises and reducing the costs associated with setting up several production lines.

[0060] When a chitosan containing a particular glucan level is sought, a first quantity of the liquid phase containing the solubilized fungal chitosan is filtered while a second quantity of the liquid phase containing the solubilized fungal chitosan is directly transferred into the precipitation tank without being filtered.

[0061] The volume of the liquid phase containing the solubilized fungal chitosan that must be derived (and therefore not undergo the optional filtration step) is determined according to the percentage of glucans desired in the final chitosan. The quantity of liquid phase containing the solubilized fungal chitosan that is derived allows the glucan level to be varied precisely and with great flexibility, for example by increasing it, depending on the intended applications for said fungal chitosan.

[0062] According to the invention, the glucan content (predetermined value depending on the intended application for chitosan). can be in a wide range of values. For example, the glucan content can be in a range between 0.1 and 50%. Preferably, the glucan content is at least 0.5%, preferably at least 1%, preferably at least 2%, preferably at least 3%, preferably at least 4%, preferably at least 5%. Preferably, the glucan content is less than 45%, preferably less than 40%, preferably less than 35%, preferably less than 30%, preferably less than 25%, preferably less than 20%. Depending on the desired application of chitosan, the required glucan level will be different and can be modulated in a flexible, controlled and precise manner using the method of the invention.

[0063] At the end of the precipitation step and as illustrated in Fig. 1, the method of the invention comprises a step of collecting said precipitated fungal chitosan. The precipitated fungal chitosan can then be washed, and / or centrifuged, and / or dried.

[0064] The process of the invention thus makes it possible to obtain a fungal chitosan derived from a mycelium characterized by a degree of acetylation of between 0 and 25%, preferably between 0 and 20%, even more preferably equal to 15%, in that it has a low molecular mass, which is expressed by a viscosity of between 1 and 15 cPs, preferably between 1 and 10 cPs, preferably between 2 and 6 cPs, and by a mass of total heavy metals < 20 mg per kg of chitosan and having a glucan level of between 0.1% and 50%.

[0065] Advantageously, the glucan level is controlled so that the glucan level is within a range of the predetermined value between 0.1% and 50%. Indeed, the glucan level can vary within a range between 0.1 and 50%. Preferably, the glucan level is at least 0.5%, preferably at least 1%, preferably at least 2%, preferably at least 3%, preferably at least 4%, preferably at least 5%. Preferably, the glucan level is less than 45%, preferably less than 40%, preferably less than 35%, preferably less than 30%, preferably less than 25%, preferably less than 20%.

[0066] Advantageously, the present invention therefore makes it possible to obtain a high added value and high quality chitosan. The process is reproducible, predictable and robust and can be used to manufacture chitosans for different applications thanks to the possibility of precisely controlling the glucan level depending on the intended application and this, for a wide range of glucan levels.

[0067] Preferably, the fungal chitosan from a mycelium obtained according to the method of the invention is in the field of health or cosmetics or oenology or agriculture or horticulture.

[0068] Advantageously, the fungal chitosan from a mycelium obtained according to the method of the invention is used by ingestion.

[0069] In a preferred embodiment, the fungal chitosan derived from a mycelium obtained according to the method of the invention is used in a composition, in particular in a pharmaceutical composition, a food supplement or a biostimulant composition.

[0070] Advantageously, the fungal chitosan derived from a mycelium obtained according to the process of the invention is used in a composition, in particular in the field of health or cosmetics or oenology or agriculture or horticulture.

[0071] Other characteristics, details and advantages of the invention will emerge from the example of the invention given below, without limitation.

[0072] Example

[0073] The mycelium and an alkaline material are fed into a 4m cylindrical-conical reactor 3equipped with a double jacket that heats the reaction medium up to 120°C. The mycelium is derived from Aspergillus niger. The alkaline material is in the form of a sodium hydroxide solution at a concentration of 50%, which is added to the cylindro-conical reactor until a pH greater than 9 is obtained in the reactor. This contacting of the mycelium with the alkaline material is carried out without adding water or solvent. Then, the mixture is stirred in the cylindro-conical reactor by a worm screw mounted on an orbital arm that travels around the periphery of the reactor at a rate of approximately 2 revolutions / min. The reactor homogenizes the mixture of alkaline material and mycelium, gradually increasing the temperature to allow the deacetylation of the chitin from the mycelium into chitosan. Deacetylation is therefore carried out at a temperature of approximately 105°C for approximately 6 hours in the cylindrical-conical reactor.Deacetylation is partial or complete with the production of chitosan in kneaded form in the form of a paste, presenting a degree of acetylation, that is to say a molar proportion of N-acetyl-D-glucosamine units along the chitosan chains, of 0 to 25%.

[0074] The resulting paste is transferred to a washing tank to be washed with water to remove the [3-glucans] during 4 successive washes in the same washing tank, then transferred to a solubilization tank. The paste is then stirred in a conventional manner in the solubilization tank for 8 hours. While stirring the paste, an acetic acid solution is added to the solubilization tank until a pH of less than 6 is obtained. At the end of solubilization, the solution contains a liquid phase containing the chitosan and an insoluble fraction containing the glucans.

[0075] After solubilization, in a directional control step, a 99% volume fraction of the liquid phase is directed to filtration and the remaining 1% volume fraction is directed to precipitation using a three-way control valve. The first way directs the liquid phase to filtration and the second way directs the liquid phase to precipitation. The volume of the liquid phase containing the solubilized fungal chitosan that is derived was determined according to the desired percentage of glucans. The amount of liquid phase containing the solubilized fungal chitosan that is derived allows the glucan cough to be modulated to the predetermined value and in a precise manner.

[0076] For the liquid phase fraction directed to filtration, the liquid phase is filtered through a cellulose plate to remove some of the remaining glucan fraction. Filtration can be followed by a turbidity measurement to assess the quality of filtration. Turbidity measurement is a measure of the cloudiness of liquids, recognized as a simple and basic indicator of liquid quality.

[0077] Then, the liquid phase fraction diverted directly to precipitation and the liquid fraction having undergone the filtration step are brought into a precipitation tank. The liquid phase is then stirred in the precipitation tank by adding sodium hydroxide until a pH greater than 9 is obtained. At a pH greater than 9, chitosan precipitates.

[0078] After precipitation, fungal chitosan is collected at the end of the process. Fungal chitosan has an acetylation degree of 0 to 25%, a low molecular weight expressed by a viscosity of 1 to 15 cPs and a total heavy metal mass of < 20 mg per kg of chitosan and a glucan content of 0.1 to 50%.

[0079] It is understood that the present invention is in no way limited to the embodiments described above and that many modifications may be made thereto without departing from the scope of the appended claims.

Claims

“DEMANDS” 1. A process for producing fungal chitosone from a mycelium comprising the following steps in order: (i) feeding a conical reactor with said mycelium and an alkaline material; (ii) at least partial deacetylation of said mycelium in said conical reactor with obtaining a kneaded fungal chitosan; (iii) at least one washing of said fungal chitosan kneaded in at least one washing vat; (iv) solubilization of the kneaded fungal chitosan in at least one solubilization tank with the obtaining of a liquid phase containing the solubilized fungal chitosan; (v) optionally a filtration of said liquid phase; (vi) precipitation of said liquid phase in at least one precipitation tank with obtaining precipitated fungal chitosan and (vii) a collection of said precipitated fungal chitosan, characterized in that it further comprises, after the solubilization step, a directional control step of said liquid phase using bypass means, during which said liquid phase is directed to said filtration and / or to said precipitation.

2. A process for producing fungal chitosan according to claim 1, wherein said at least partial deacetylation of the mycelium is carried out at a temperature between 100 and 110°C, preferably between 103 and 107°C, and even more preferably at a temperature of about 105°C.

3. A process for producing fungal chitosone according to any one of the preceding claims, wherein said at least partial deacetylation of the mycelium is carried out for a period of between 4 hours and 10 hours, and preferably for about 6 hours.

4. A process for producing fungal chitosan according to any one of the preceding claims, wherein said alkaline material is in the form of an alkaline solution selected from an aqueous solution of sodium or potassium hydroxide or a mixture thereof, and is preferably an aqueous solution of sodium hydroxide.

5. A process for producing fungal chitosan according to any one of the preceding claims, wherein said alkaline solution has a concentration of between 40% and 60%, preferably between 45% and 55%, even more preferably 50%.

6. A process for producing fungal chitosan according to any one of the preceding claims, wherein the addition of the alkaline material is carried out until a pH adjusted to a value greater than 9.0, preferably greater than 9.5, and even more preferably adjusted to 10.

7. A process for producing fungal chitosan according to any one of the preceding claims, wherein said solubilization of the kneaded fungal chitosan is carried out by adding an acid in the form of an acidic solution until a pH of less than 6 is obtained, preferably less than 5.5 and even more preferably less than 4.

5.

8. A process for producing fungal chitosan according to any one of the preceding claims, wherein the solubilization of the kneaded fungal chitosan is carried out during a duration between 7 and 9 hours, preferably around 8 hours.

9. A process for producing fungal chitosan according to claim 7 or 8, wherein said acid is selected from the group comprising hydrochloric acid, lactic acid, formic acid, succinic acid, glycolic acid, citric acid, acetic acid and mixtures thereof.

10. Process for producing fungal chitosan according to any one of claims 7 to 9, wherein said acid solution has a concentration between 0.1 and 1 N.

1. A process for producing fungal chitosan according to any one of the preceding claims, wherein, during said directional control step, at least 0.1% by volume of said liquid phase, preferably at least 0.5% by volume of said liquid phase, preferably at least 1% by volume of said liquid phase, preferably at least 1.5% by volume of said liquid phase, preferably at least 2% by volume of said liquid phase, is directed towards said precipitation.

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