Chitin and Chitosan

Monday, 02 May 2005 00:00 Dr. Beaulieu
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ImageOnce words such as chitin and chitosan are in your mental radar screen, you will find them everywhere. Possibly, you had a lobster supper last night and you removed that hard outer shell as a useless stuff. In fact, you discarded the hard shell that consisted millions of tightly interwoven polymer strands called chitin. The hard outer shell, or exoskeleton, are known to give protection to shrimps, crabs, lobsters, scorpions, insects etc. from their predators.Chitin is one of the most abundant polysaccharide in nature1, being only second after cellulose. It can be found in animals (exoskeletons of crustacean and insects) as well as in fungi, mushrooms and yeasts2.

A brief background of chitin

The basic principles of the chitin isolation are known since the beginning of 19th century. It was, Professor Henri Braconnot, Director of the Botanical Garden in Nancy, France first isolated a fraction called fungine in 1811 from the cell walls of mushrooms. In 1823 Odier renamed fungine as chitin (meaning tunic in Greek) almost 3 decades before the isolation of cellulose. Chitin is mostly obtained from the exoskeleton of industrially processed crustaceans, such as lobster, crab and shrimp which contains between 20 to 40% of chitin3. The increased use of chitin (and its derivates) is motivated by the fact that contrary to the petroleum derivatives, chitin is obtained from fisheries by-products, naturally renewable source, non-toxic, non-allergenic, anti-microbial and biodegradable.



Chemistry of chitin

Chitin is a polysaccharide. A polysaccharide is a polymer - a giant molecule consisting of smaller molecules of sugar strung together. Chitin can be described as a biopolymer composed of N-acetyl-D-glucosamine; a chemical structure very close to cellulose except that the hydroxyl group in C (2) of cellulose being replaced by an acetamido group in chitin. One can associate this chemical similarity between cellulose and chitin as serving similar structural and defensive functions2.

 

How to extract chitin from the crustacean (hard) shells?

While there exists many extraction methods of the chitin from the crustacean shells, the principles of chitin extraction are relatively simple. The proteins are removed by a treatment in a dilute solution of sodium hydroxide (1-10%) at high temperature (85-100°C). Shells are then demineralized to remove calcium carbonate. This is done by treating in a dilute solution of hydrochloric acid (1-10%) at room temperature. Depending on the severity of these treatments such as temperature, duration, concentration of the chemicals, concentration and size of the crushed shells, the physico-chemical characteristics of the extracted chitin will vary. For instance, the three most important characteristics of the chitin i.e., degree of polymerization, acetylation and purity, will be affected. Shell also contains lipids and pigments. Therefore, a decolorizing step is sometimes needed to obtain a white chitin. This is done by soaking in organic solvents or in a very dilute solution of sodium hypochlorite. Again, these treatments will influence the characteristics of the chitin molecule.



Chitosan - another important derivative of chitin

Chemists love to play with molecules. They did not spare chitin, the polymer either and made chitosan. The term chitosan is used when chitin could be dissolved in weak acid. When chitin is heated in a strong solution of sodium hydroxide (>40%) at high temperature (90-120°C), chitosan is formed. This harsh treatment removes acetylic grouping on the amine radicals to a product (chitosan) that could be dissolved. It is said that at least 65% of the acetylic groups should be removed on each monomeric chitin to obtain the ability of being put in solution2. The degree of deacetylation will vary according to the duration, the temperature and the concentration of the sodium hydroxide. Furthermore, many chemical characteristics of the chitosan (molecular weight, its polydispersity, the purity) are greatly dependant on the method, the equipment used and also of the source of the shells. It is therefore, crucial to control precisely methods of production of the chitosan to obtain the exact characteristics needed for end use application of the product.

 

Making chitosan into a value-added ingredient - how?

Three main characteristics of chitosan to be considered are: molecular weight, degree of deacetylation and purity. Since chitosan is a polymer formed by repeating units of D-glucosamine (sugar), the total length of the molecule is an important characteristic of the molecule. As a result, the molecular weight is a key feature for a particular application of chitosan. The molecular weight of the native chitin has been reported to be as high as many million Daltons. However, the harsh chemical treatment tends to bring down the molecular weight of the chitosan, ranging from 100 KDa to 1500 KDa. An inert environment during the deacetylation could preserve the molecular chain. On the other hand, low molecular weight could be produced by different ways including enzymatic or chemical methods. When the chain becomes short to shorter, chitosan could be dissolved directly in water without the need of an acid. This is particularly useful for specific application in cosmetic or in medicine when the pH should stay around 7.0. Molecular weight of chitosan could be measured by gel permeation chromatography, light scattering, or viscometry. Because of simplicity, viscometry is the most commonly used methods even though it is influenced by many factors (concentration temperature, ionic strength, pH, type of acids) other than the molecular weight.
Since chitosan is made by deacetylation of chitin, the term degree of deacetylation (DAC) is used to characterize chitosan. This value gives the proportion of monomeric units of which the acetylic groups that have been removed, indicating the proportion of free amino groups (reactive after dissolution in weak acid) on the polymer. DAC could vary from 70 to 100%, depending of the manufacturing method used. This parameter is important since it indicates the cationic charge of the molecule after dissolution in a weak acid. There are many methods of DAC measurements like UV and infrared spectroscopy, acid-base titration, nuclear magnetic resonance, dye absorption, etc.. Since there are no official standard methods, numbers tend to be different for different methods. In high value product, NMR gives the most precise DAC number. Given its high cost, many producer uses titration or dye adsorption as a quick and convenient method, that yields similar results as NMR.
Finally, the purity of the product is vital particularly for high-value product (biomedical or cosmetic area). This purity is quantified as the remaining ashes, proteins, insolubles, and also in the bio-burden (microbes, yeasts and moulds, endotoxins). Even in the lower value chitosan such as that used for the wastewaters treatment, the purity is a factor because the remaining ashes or proteins tend to block active sites, the amine grouping. Being not available to bind, a greater amount of chitosan is needed to be effective.

 

Here comes the multitude of applications …

Chitosan is a biological product with cationic (positive electrical charge) properties. It is of great interest, all the more so because most polysaccharides of the same types are neutral or negatively charged. By controlling the molecular weight, the degree of deacetylation and purity, it is possible to produce a broad range of chitosans and derivatives that can be used for industrial, dietary, cosmetic and biomedical purposes. Together these properties have led to the development of hundreds of applications so far. There are plethora of literature, books and conference proceedings that documented the multiple uses of the chitosan4. It is out of the scope of this article to describe extensively every applications of chitosan. We will concentrate on the major uses of chitosan and the most promising future applications. Applications of chitosan can be classified mainly in 3 categories according to the requirement on the purity of the chitosan:



In agriculture:

Chitosan offers a natural alternative to the use of chemical products that are sometimes harmful to humans and their environment. Chitosan triggers the defensive mechanisms in plants (acting much like a vaccine in humans), stimulates growth and induces certain enzymes (synthesis of phytoalexins, chitinases, pectinases, glucanases, and lignin). This new organic control approach offers promise as a biocontrol tool. In addition to the growth-stimulation properties and fungi, chitosans are used for:

 

For water treatment:

At the present time, physicochemical-type treatment is widely used at potable and wastewater treatment plants. The major disadvantage of using synthetic chemical products is the risk of resulting environmental pollution. Treating wastewater using "greener" methods has become an ecological necessity. Chitosan, due to its natural origin and being biodegradable, has proven to be a most interesting alternative from several points of view.
Integrating a natural polymer made of crustacean residue into an existing system achieves a two-fold purpose: it improves the effectiveness of water treatment while reducing or even eliminating synthetic chemical products such as aluminum sulphate and synthetic polymers. Here are a few characteristics of chitosan that offer an ecological solution:

 

 

In food:

Chitosan is already used as a food ingredient in Japan, in Europe and in the United States as a lipid trap, an important dietetic breakthrough. Since chitosan is not digested by the human body, it acts as a fiber, a crucial diet component. It has the unique property of being able to bind lipids arriving in the intestine, thereby reducing by 20 to 30% the amount of cholesterol absorbed by the human body. This raises the question: is chitosan really a "Fat Magnet"?
In solutions, chitosan has thickening and stabilizing properties, both essential to the preparation of sauces and other culinary dishes that hold their consistency well. Finally, as a flocculating agent, it is used to clarify beverages. Because of its phytosanitary properties, it can be sprayed in dilute form on foods such as fruits and vegetables, creating a protective, antibacterial, fungi static film. In Japan, a dilute solution of chitosan is commonly sprayed on apples and oranges as a protective measure. There are many other applications in the areas of nutraceutical and nutritional supplements, particularly for the broad range of chitosans that have been chemically or enzymatically modified.
Principal commercial applications include:

 

In cosmetics:

Chitosan forms a protective, moisturizing, elastic film on the surface of the skin that has the ability to bind other ingredients that act on the skin. In this way, chitosan can be used in formulating moisturizing agents such as sunscreens, organic acids, etc. to enhance their bioactivity and effectiveness. Today, chitosan is an essential component in skin-care creams, shampoos, and hairsprays due to its antibacterial properties. Many patents have been registered and new applications are just beginning to appear including the most highly prized moisturizing and antibacterial properties. Applications include

 

For biopharmaceutical uses:

It is in the field of health that the many properties of chitosan (bacteriostatic, immunologic, antitumoral, cicatrizant, hemostatic and anticoagulant) are of interest. For example, because of its biocompatibility with human tissue, chitosan's cicatrizant properties have proven its effectiveness as a component, notably, in all types of dressings (artificial skin, corneal dressings, etc.), surgical sutures, dental implants, and in rebuilding bones and gums. Applications currently being developed include artificial skin, surgical sutures that are absorbed naturally after an operation, and corneal contact lenses. Finally, chitosan delivers and time-releases drugs used to treat animals and humans. There are many potential chitosan applications in the health field but their development calls for the use of components that comply with strict pharmaceutical-grade requirements.
Possible applications include:

 

Looking forward ...

Applications of chitosan is growing rapidly. Not only due to its multitude of applications but due to increasing environmental awareness of the population, biodegradable, and non-toxic products from 'natural' sources such as chitin and chitosan are going to be more and more appealing for the replacement of synthetic compounds. Moreover, in cosmetic and in biopharmaceutical industries, chitosan has exclusive properties which are not found in other synthetic products.

 

References

1. Knorr, D. Functional properties of chitin and chitosan. J. Food Sci. 47, pp. 593-595, 1982.
2. Roberts, G.A.F. Chitin Chemistry, Macmillan, London, 1992.
3. No, H.K. and Meyers, S.P. (1995). Preparation and characterization of chitin and chitosan (A review). J. Aquatic Food product Technol., 4, pp. 27-52 (1995).
4. Gossen, M.F.A. (1997) Applications of Chitin and Chitosan. Technomic Publishing Company Book, Lancaster, 1997.

 

Clermont Beaulieu

Clermont Beaulieu, Marinard Biotech Inc., 30 de l'Entrepôt, Rivière-au-Renard (Qc),G4X 5L4 Canada



Clermont Beaulieu received his Ph.D. from Laval University (Québec City), and did his post-doctoral studies at the Dalhousie University (Halifax) and at the University of Oxford (England). He then joined University of British Columbia as an Assistant Professor in 1991. Moved back to University of Montreal as a Professor (1991-1997) at the department of pathology. Currently, he is Asssociate Professsor at the Dept. of Biology, Université du Québec à Rimouski since 2001.



Dr. Beaulieu's research focuses on value-added products from seafood processing wastes as he continues to develop production facilities (industrial, food-grade and pharmaceutical grade) for extracting value-added bio-molecules from the sea. He has authored over 80 papers and Chaired conferences. His many honors and prizes include scholar of the Canadian Medical Research Council & Fonds de la recherche en santé du Québec (1982-'97), Quebec annual forum of chemical industries (2001), Canada Economic Development for the excellence of the regional project, and from the regional exportation bureau for business growth (2004).



Currently, he is the CEO of Marinard Biotech.