The Most Common Biochemicals On This Planet: Chitin

Editor’s Note: Whole dried insects are about 10 percent chitin, more or less. Although chitin presents problems of digestibility and assimilability in monogastric animals, it and its derivatives, particularly chitosan, possess properties that are of increasing interest in medicine, industry and agriculture. If the time should come when protein concentrates from insects are acceptable and produced on a large scale, the chitin byproduct could be of significant value. At the editor’s request Dr. Walter G. Goodman. professor of developmental biology in the Department of Entomology, University of Wisconsin, kindly agreed to prepare a short article for the Newsletter on the characteristics of chitin and some of its potential applications.

What if someone told you that one of the most common biochemicals on this planet was capable of,
-significantly reducing serum cholesterol
-acting as a hemostatic agent for tissue repair
-enhancing bum and wound healing
-acting as an anticoagulant
-protecting against certain pathogens in the blood and skin
-serving as a nonallergenic drug carrier
-providing a high tensile strength biodegradable plastic for numerous consumer goods
-enhancing pollutant removal from waste-water effluent
-improving washability and antistatic nature of textiles
-inhibiting growth of pathogenic soil fungi and nematodes
-boosting wheat, barley, oat, and pea yields as much as 20%

Preposterous? Unlikely? Investigators have recently proposed that chitin (see Figure la, page 6), a carbohydrate polymer found in invertebrate exoskeletons, protozoa, fungi, and algae is the polymer of the future, with numerous applications to agricultural, biomedical and consumer needs. Its apparent abundance, in combination with its toughness yet biodegradable properties, has made chitin and its derivatives the focus of interest for scientists around the world.

Chitin, an unbranched, poly-B-(14) linked N-acetyl-D-glucosamine, is present in long chains that are often oriented so that adjacent chains are antiparallel. In insects, chitin occurs as a collection of microfibrils approximately 25Ain diameter. These microfibrils form a crystalline array and are ensheathed in a matrix of cross-linkng protein. The tissue primarily involved in the synthesis of chitin, the epidermis, is a single layer of cells that secrete the chitin microfibrils so as to form layers of chitin that are parallel to the upper surface of the cell. As the process continues, the newer layers are secreted in a parallel fashion but the orientation of the fibrils has been slightly rotated. This can be best visualized by placing one hand over the other in a parallel fashion and outstretching the fingers. Keeping the bottom hand in the same plane, rotate it slightly so that looking from the top side the fingers form a grid. As your hands rotate through 180°, note the spatial orientation between the fingers of the upper and lower hands. On a smaller scale, a similar process occurs in the newly secreted layers of fibrils. Thus, micrograph cross-section through the cuticle resembles the end-view of plywood due to the varied orientation of the parallel layers of microfibrils.

Global bioproduction of chitin is enormous and estimated to be greater than 109 tons per year. Such numbers provide the entrepreneurial mind with thoughts of untold quantities of inexpensive, raw product. Yet in a practical sense, the abun-dance of exploitable chitin is severely limited. Although insects and fungi have the highest ratio of chitin to body mass, the primary source has been the shellfish industry. Waste products generated from crab and shrimp processors represent a reliable but rather limited source of chitin, thus restricting the use of chitin and its derivatives primarily to high value-in-use utilizations. Moreover, the seasonability of the supply, variability in product quality, and scattered distribution points reduce the attractiveness of chitin as a useful biopolymer. Coupled with these restrictions, chitin is insoluble in both water and most common organic solvents which makes its use in the production of fibers, membranes or agricultural products difficult. Accordingly, considerable emphasis has been placed on the dissolution and restructuring of the matrix and on the formation of soluble chitin derivatives.

One of the most useful derivatives of chitin has been chitosan. This partially deacetylated chitin, discovered in the last century, is produced by boiling chitin in concentrated base. Although chitosan is naturally occurring in certain fungi and green algae, its commercial source remains the shellfish industry. Crab and shrimp exoskeletons are pulverized, then treated with sodium hydroxide to solubilize the protein matrix surrounding the microfibrils and to dissolve residual cellular debris adhering to the carapace. The flakes are then washed and demineralized with hydrochloric acid. Crustacean exoskeleton, in contrast to that of most insects, contains calcium, as much as 20%, and this must be removed to yield pure chitin. The purified chitin is then deacetylated by treatment in concentrated sodium hydroxide to produce chitosan (see Figure lb, page 6). Unfortunately, bulk production of chitin and chitosan in this manner leads to chain breakage and unpredictable alteration of the component sugars. Improved methods are now available that eliminate these problems.

In contrast to chitin, chitosan is soluble in dilute acidic solutions and is the major outlet for chitinous products. Once in solution, it can then be fabricated into gels, films, and powders and then modified by various chemical processes including acylation, aldimination, carboxymethylation, phosphorylation and sulfation to form the desired products.

Chitin and its derivatives have been used in a number of ways. Historically, chitin was first used in wound healing. Koreans have long used the pen of an octopus as a source of chitin for the treatment of abrasions, while Mexicans have used mushrooms with their chitosanaceous cell walls to accelerate laceration wound healing. Advances in biotechnology have capitalized on these observations to provide potentially new approaches to medical problems. For example, chitosan has been used experimentally in the treatment of burns because it forms a tough, water-absorbent, biocompatible film that prevents bacterial invasion. This film can be applied directly to the burn by application of an aqueous solution of chitosan acetate. Since the underivatized chitosan is slowly degraded by the enzyme lysozyme, periodic removal of the film is unnecessary. The treatment of grafting material used to stop life-threatening vascular bleeding with chitosan has also yielded promising results. Hemostatic graft material was treated with chitosan acetate and applied to experimentally induced vascular wounds. Chitosan-treated grafts performed as well as the routinely used blood-treated grafts. In some whole animal studies, chitosan-treated grafts inhibited fibroplasia and stimulated the regeneration of normal tissue elements.

Bioerosion of chitin derivatives offers new approaches to controlled delivery of drugs. For instance, if therapeutic agents are required on a long-term basis, a single implantation of capsules containing the agent admixed with the chitin derivative could serve as gradual release sites. As enzymes hydrolyze the matrix, the drug is slowly released. Another potential use for chitin derivatives is in the production of contact lenses. A lens composed of chitosan derivatives offers the attractive properties of being tough, highly moldable, transparent, water absorb-ent and oxygen permeable. These properties make chitosan-based soft contact lenses potentially useful for extended wear. Limited studies have now demonstrated that chitosan can ameliorate dermatitis in monkeys and humans and can stimulate in a nonspecific manner the immune system of rodents. Sulfated N-carboxymethyl derivatives of chitosan have been demonstrated to block blood clotting in vitro. This is not surprising considering that the potent anticoagulant, heparin, is a highly sulfated polysaccharide with a chemical structure not unlike that of the derivatized chitosan.

Figure 1: Structure of chitin and one of its derivatives, chitosan. Both compounds are polymers of varying lengths.

In a cholesterol conscious society, the discovery of low toxicity drugs that lower serum cholesterol levels is welcome. Although eating raw insects won’t change your serum cholesterol levels significantly, the use of certain chitin derivatives may. A number of studies have demonstrated that rats fed a diet containing 2-5% chitosan can substantially reduce serum cholesterol with few side effects. Chitosans are thought to: (1) bind bile acid and/ or cholesterol in the intestinal lumen, thus reducing uptake, and (2) stimulate excretion by interfering with the absorption process. Thus, an inexpensive, nonnutritive agent such as chitosan, capable of inducing hypocholesterolemic activity should be competitive with the more expensive drugs such as cholestyramine.

From an industrial viewpoint, the interaction of chitin derivatives with various toxic metals provides a means of removing pollutants from industrial waste water. Interestingly, derivatives of cellulose, normally used for such purposes, have significantly less capacity for ion binding than does the chitin derivative. On the other hand, the hydrophobic nature of the fibrils makes them less likely to swell, thus permitting high throughput of aqueous waste effluent.

The extensive use of non-biodegradable plastic in modem societies has generated a waste disposal crisis as well as problems for marine organisms. This point is underscored by the estimate that 30% of the world’s ocean fish have tiny pieces of plastic in their stomachs that interfere with digestion. Much of the environmental damage attributable to plastics is caused by precisely those characteristics for which they were developed, namely, their durability over products constructed from natural materials. As noted earlier, chitin derivatives can be cast into filaments and films of high tensile strength owing to their extensive hydrogen bonding in three dimensions. This degree of cross-linking makes them much stronger than cellulose. Despite its strength, materials derived from chitosan can be rapidly degraded by soil or marine microorganisms. Indeed, when chitosan films constructed of the appropriate chainlength were incubated in cold ocean waters, complete decay was observed within a month. Although chitosan represents a promising approach to the problem of biodegradability, the limited supplies of raw product cannot begin to supply the voracious demand for plastics. Until massive sources of chitin become readily available, dependence upon petrochemically-derived polymers remains the reality.

The use of chitin-based derivatives is most widespread in agricultural applications. One west coast company offers chitosan as a seed treatment for the control of nematodes and fungi in wheat, barley, oats and peas. The mechanism by which chitosan suppresses fungi and nematodes is uncertain although several hypotheses have been presented. Unforunately, the amount of chitosan required to effect a general reduction in soil pathogens is enormous and the nematocidal levels of chitin may be phytotoxic. Added to this is the problem of variability between chitosan preparations. The source of chitin seems to play an unknown role in the effectiveness as a fungicide or nematocide. Although full-scale agricultural application of chitin-based derivatives is unlikely in the near future, this does not dampen the outlook for utilization of chitin in agriculture. For example, in areas where large-scale insect outbreaks occur, insect bodies might be used as a source of fertilizer as well as a source of chitin. Better methods may be forthcoming in reducing the variability in chitosan preparation and specific fractions of chitosan may be identified as to the active ingredient.

With the many possibilities that chitin-based products offer given the means of mass production and genetic engineering, the future of this biopolymer is bright. For further information about chitin, chitosan, and the many derivatives, see below:

Bade, M.L.; Wick, R.L. 1988. Protecting crops and wildlife with chitin and chitosan. In Biologically Active Natural Products, edited by H.G. Cutler, American Chemical Society, Washington, D.C. pp. 450-468.
Hepburn, H.R. 1976. The Insect Integument, Elsevier Scientific, Amsterdam.
Kramer, K.J.; Dziadik-Turner, C.; and Koga, D. 1985. Chitin metabolism in insects. In Comprehensive Insect Physiology, Biochemistry and Pharmacology, Pergamon Press, Oxford. pp. 75-115.
Muzzarelli, R.; Jeuniaux, C.; Gooday, G.W. 1986. Chitin in Nature and Technology, Plenum Press, N.Y.
Neville, A.C. 1975. Biology of the Arthropod Cuticle, Springer-Verlag, N.Y.
Zikakis, J.P. 1984. Chitin, Chitosan, and Related Enzymes,
Academic Press, N.Y.

by Dr. W.G. Goodman (University of Wisconsin-Madison)