WO2008007321A2

WO2008007321A2 - Cross-linked chitosan and process to cross-link

Info

Publication number: WO2008007321A2
Application number: PCT/IB2007/052659
Authority: WO
Inventors: Cosmo Mezzina; Giovanni Scapagnini; Ivo Volpato; Bernard Bizzini; Giovanni Franchi
Original assignee: Sirc SpA Natural & Dietetic Foods
Current assignee: Sirc SpA Natural & Dietetic Foods
Priority date: 2006-07-10
Filing date: 2007-07-06
Publication date: 2008-01-17
Anticipated expiration: 2009-01-10
Also published as: ITMI20061335A1; EP2044127A2; WO2008007321A3

Abstract

The present invention relates to Chitosans in crosslinked form wherein the crosslink derives from the reaction of said chitosans with a crosslinker chosen from among the group of aliphatic aldehydes, and to a Chitosan crosslinking process comprising the following stages: 1) suspend chitosan in a buffer solution having a pH that ranges between 7.5 and 9.5, preferably between 8.0 and 9.0, and more preferably between 8.1 and 8.3 (suspension 1); 2) place suspension (1) in contact with an aliphatic aldehyde as crosslinker and leave to react for 1 to 24 hours at a temperature of between 15- and 40 -C (suspension 2); and possibly: 3) treat suspension (2) with an organic amine compound.

Description

[01] This invention relates to chitosans, which are polymers of D-glucosamine, in crosslinked form, and to a process for producing chitosans in crosslinked form.

[02] Said polymers are a basic component of some diet supplements, possibly in association with other compounds, used to produce cholesterol-lowering and lipid-lowering effects and to reduce sugar absorption and body weight, as described, for example, in Italian patent 1284638 filed by the Applicant.

[03] More specifically, from the chemical standpoint, the chitosans are poly(β- 1 ,4-glucosamine).

[04] It has now surprisingly been found that the chitosans are crosslinkable. The crosslinking reaction involves the free amine functional groups present in the macromolecule of the chitosans. These functional groups react with a suitable crosslinker, forming stable bonds with them. In particular, chitosans cross-linked according to this invention have crosslinking bonds which are irreversible under the physiological conditions of the human body. The crosslinked chitosans according to this invention, compared with the non-crosslinked form, therefore cannot be attacked or degraded (or are attacked or degraded to a much lesser extent) by digestive enzymes and/or the physiological conditions of the gastrointestinal tract, such as the pH. As a result, said chitosans pass through the entire digestive apparatus practically unchanged.

[05] This invention consequently relates to chitosans in crosslinked form wherein the crosslink derives from the reaction of said polymer with a crosslinker chosen from among the group of aliphatic aldehydes.

[06] The crosslinked chitosans according to this invention can have a very broad range of crosslinking, from totally cross-linked to very slight crosslinking. In this invention, "degree of crosslinking" means the ratio between the number of functional amine groups (-NH2) which have reacted with a functional aldehyde group of the crosslinker and the total number of functional amine groups present in the polymer.

[07] The crosslinked chitosans according to the invention have a degree of crosslinking which varies between approximately 1 :10 and 1 :200, preferably between 1 :30 and 1 :150, and more preferably between 1 :60 and 1 :120.

[08] The aliphatic aldehyde is preferably a hydrocarbon with a C1-C10 alky] chain. The aldehyde is preferably chosen from among the monoaldehydes and dialdehydes. The preferred example of monoaldehyde is formaldehyde. The particularly preferred example of dialdehydes is glutaraldehyde ((OHC-(CH₂)S-CHO)).

[09] Any chitosan can be crosslinked. The crosslinked chitosans used according to this invention have an apparent density which ranges between approximately 0.3 and 0.80 g/ml, and preferably between 0.5 and 0.7 g/ml. The particle size of these polymers typically ranges between 80 and 400 mesh, and preferably between 80 and 20 mesh. The chitosans have a degree of deacetylation which typically ranges between 89 and 95%, but those with a degree of deacetylation exceeding 90% are preferred.

[010] The crosslinking process, which is a further subject of this invention, comprises the following stages:

[011] 1) suspend chitosan in a buffer solution having a pH that ranges between 7.5 and 9.5, preferably between 8.0 and 9.0, and more preferably between 8.1 and 8.3 (suspension 1);

[012] 2) place suspension (1) in contact with an aliphatic aldehyde as crosslinker and leave to react for 1 to 24 hours, preferably 1 to 18 hours, and more preferably 1 to 2 hours, at a temperature of between 15° and 40°C, and preferably between 20 and 27°C (suspension 2); and possibly:

[013] 3) treat suspension (2) with an organic amine compound.

[014] Said buffer solution is obtained by adding acid salts of alkaline metals to the water, preferably a pair of sodium acid phosphates, namely Na₂HPO₄ and NaH₂PO₄, or the sodium carbonate/bicarbonate pair.

[015] Said buffer solution has a salt molarity of between approximately 0.01 and 0.5, and preferably between 0.05 and 0.1.

[016] The quantity of aliphatic aldehyde reacted with the polymer to be crosslinked is chosen on the basis of the degree of crosslinking to be obtained and the number of functional aldehyde groups in the molecule. The ratio between chitosan and crosslinker, expressed as moles of glucose contained in the polymer compared with the moles of crosslinker, typically ranges between 1 :1 -1CM and 1 :1 -10-³.

[017] For example, if 100 g of chitosan (corresponding to 6.17 x 10"¹ of glucose) is reacted with 6.17 x 10-² moles of glutaraldehyde, a crosslinking ratio of 1:10 will be obtained, while a crosslinking ratio of 1 :200 is obtained by reacting 100 g of chitosan with 3.085 x 10'³ moles of glutaraldehyde.

[018] The crosslinker can be added, for example, in the form of an aqueous solution. In said solution the molarity of the crosslinker, namely the aldehyde, can range, for example, between 0.1 and 10, and preferably between 0.5 and 2.

[019] Said organic amine compound is preferably an aminoacid, such as glycine or lysine. Said amine compound is added greatly in excess of the aldehyde functions which have not yet reacted; it is generally a solution which has approximately the same molarity as the solution containing the aldehyde.

[020] The suspension is preferably stirred at stages (2) and (3).

[021] Non-limiting examples of this invention are set out below. The following test methods were used to define this invention:

- degree of crosslinking: the -N Hb functions still free are titrated with Habeeb's dinitrobenzene sulphonate (DNBS) method (reference A.G.S.A. Anal. Biochim., 1966, 14, 328-336). The number of -NH2 functions in the titrated polymer being known, the degree of crosslinking is calculated;

- apparent density: measured using an ERWEKA SVM 202 densitometer, SERIAL NUMBER 113260.03ec, with the following method:

I. place a known quantity in weight of the powder to be tested in the graduated cylinder of the instrument; 1. set the number of strokes of the instrument to 250; [022] III. start the instrument;

1. read the volume occupied by the powder on the graduated cylinder; and 2. calculate the apparent density of the powder under test with the following formula:

[023] APPARENT DENSITY = MASS / VOLUME = g / ml [024] Characteristics of the polymers used in the examples:

- chitosan: apparent density 0.6 g/ml, particle size 100 mesh and degree of deacetylation 95%, and

- chitin: apparent density 0.6 g/ml and particle size 100 mesh. [025] Example 1

[026] Introduce 10 kg of chitosan into an apparatus equipped with a stirrer and pour 100 I of an aqueous solution of 0.07 M Na2HPO₄ at pH 8.2 onto it. Begin stirring and add 1 I of an aqueous solution of 1 M glutaraldehyde to the suspension thus obtained. Leave to react at ambient temperature for 6 hours, maintaining stirring. Stop the reaction by adding excess glycine (1 I) and filter the suspension through a Bϋckner filter to separate the chitosan from the reaction liquid; then wash with water until all trace of unreacted glutaraldehyde has been eliminated. Under said conditions, the degree of crosslinking is 1 :120.

[027] Example 2

[028] Repeat example 1 , with the sole difference that 2 I of an aqueous solution of glutaraldehyde is added instead of 1 I.

[029] A crosslinked chitosan with a degree of crosslinking of 1:60 will be obtained.

Claims

1. Chitosans in crosslinked form wherein the crosslink derives from the reaction of said chitosans with a crosslinker chosen from among the group of aliphatic aldehydes.

2. The chitosans claimed in claim 1 , wherein the aliphatic aldehyde is a hydrocarbon with a d-C-io alkyl chain.

3. The chitosans claimed in claim 2, wherein the aliphatic aldehyde is chosen from among the monoaldehydes and dialdehydes.

4. The chitosans claimed in claim 2, wherein the aliphatic aldehyde is chosen from formaldehyde and glutaraldehyde.

5. The chitosans claimed in claims 1 to 5, wherein the degree of crosslinking ranges between approximately 1:10 and 1:200.

6. The chitosans claimed in claim 5, wherein the degree of crosslinking ranges between 1 :30 and 1 :150.

7. The chitosans claimed in claims 1 to 7, wherein the chitosans to be crosslinked have an apparent density ranging between 0.3 and 0.80 g/ml, a particle size of 80 to 400 mesh and a degree of deacetylation which typically ranges between 89 and 95%.

8. The chitosans claimed in claim 7, wherein the chitosans to be crosslinked have an apparent density ranging between 0.5 and 0.7 g/ml, a particle size of 80 to 20 mesh and a degree of deacetylation exceeding 90%.

9. Chitosan crosslinking process comprising the following stages:

1) suspend chitosan in a buffer solution having a pH that ranges between 7.5 and 9.5, preferably between 8.0 and 9.0, and more preferably between 8.1 and 8.3 (suspension 1);

2) place suspension (1) in contact with an aliphatic aldehyde as crosslinker and leave to react for 1 to 24 hours at a temperature of between 15° and 40°C (suspension 2); and possibly:

3) treat suspension (2) with an organic amine compound.

10. The process claimed in claim 9, wherein the aliphatic aldehyde is a hydrocarbon with a C1-C10 alkyl chain.

11. The process claimed in claim 10, wherein the aliphatic aldehyde is chosen from among the monoaldehydes and dialdehydes.

12. The process claimed in claim 11 , wherein the aliphatic aldehyde is chosen from formaldehyde and glutaraldehyde.

13. The process claimed in claims 9 to 12, wherein the buffer solution is an aqueous solution of acid salts of alkaline metals.

14. The process claimed in claim 13, wherein the buffer solution is formed with a pair of sodium acid phosphates or a sodium carbonate/bicarbonate pair.

15. The process claimed in claims 9 to 14, wherein the ratio between chitosan and crosslinker, expressed as 1 gram of polymer to moles of crosslinker, ranges between 1 :1 -10-1 and 1 :1 -10-3.

16. The process claimed in claims 9 to 15, wherein the reaction time at stage (2) ranges between 1 and 2 hours.

17. The process claimed in claims 9 to 16, wherein the reaction temperature at stage (2) ranges between 20 and 27°C.

18. The process claimed in claims 9 to 18, wherein the chitosans to be crosslinked have an apparent density ranging between 0.3 and 0.80 g/ml, a particle size of 80 to 400 mesh and a degree of deacetylation which typically ranges between 89 and 95%.

19. The process claimed in claims 9 to 18, wherein the chitosans to be crosslinked have an apparent density ranging between 0.5 and 0.7 g/ml, a particle size of 80 to 20 mesh and a degree of deacetylation which exceeds 90%.