Production and Functionalization of Chitosan Films Fortified with Quinones As Anti-Microbial Bio-Bandages

Production and Functionalization of Chitosan Films fortified with Quinones as Anti-Microbial Bio-Bandages

Mohammad Alcheikh Ali *, Nizar Issa*

A-   The previous status of wound dressings:

Conventional bandages used in many pharmaceutical industries have lots of disadvantages caused by their composition , they are made from such materials that negatively impact on general health in addition to environmental concerns because they degrade under highly cost conditions and human efforts. Furthermore, they may be contagious and conduct infections due to their direct adhesiveness onto wounds, burns and ulcers which pollutes the bandage that does not have anti-bacterial activity and does not help in wound healing.

Since conventional bandages have above mentioned disadvantages, we have fabricated new wound dressings from organic abundant biodegradable material from marine crustaceans in Syrian fisheries and marine food restaurants such as Shrimp shells that had been used as wound dressings by ancient chineses warriors since centuries, this material is called chitosan. The chitosan have had strong anti-microbial activity, natural bio-adhesive properties, permeability to oxygen and water vapor, non-inflammatory and accelerate wound healing. Conventional bandages may isolate wounds from microbes but, my bio-bandages have anti-microbial effects, help in wound healing by accelerating wound recovery. in addition to that , they do stimulate immunological responses against pathogenic agents. Throughout the ages, Olive tree has got historical, economical and social importance in Syria and Syrian Ebla city in Idleb is considered the first home for olive tree in the world and The cultivation of olive in Syria composes 12% of the total cultivated area and 65% of the total area of fruit trees and thus constitute a rich source of natural phenoles that can be used to strengthen and improve the bio- bandages characteristics.

^{*Mohammad Alcheikh Ali: master of biotechnology student , biomaterial production and functionalization.}

^{*Nizar Isaa:prof.at the fac of sciences, univ. Damascus, researcher at national commission for biotechnology.}

We can strengthen the bio-bandages made ​​from Chitosan extracted from shrimp shell wastes by the results of enzymatic oxidation (by laccase enzyme) of phenols extracted from olive pomace yielding phenol complexes (secondary or triple) according to the phenolic substrate used which binds to chitosan molecules on its part contains of D-glucosamine groups , specifically with NH2+ groups in order to play their anti-bacterial and anti-inflammatory role to improve the dynamic characteristics of chitosan based dressings and access to the best performance in the resistance of bacteria and fungi and pathogens, which provides a solid defense and full isolation of wounds and burns for contaminants that may cause negative repercussions in the wound area and the modified bio-bandages are supposed to destroy and inhibit the work of a wider range of pathogenic microorganisms in wounds that find an appropriate place for feeding and breeding.                                               

 As phenol complexes grafted on the chitosan bind to the metal ions within the structure of bacterial enzymes such as iron hindering the metabolism of bacteria and thus contribute to the elimination of bacteria in addition to the role of phenols in reduction of the oxygen exists in the area between the bandage and the wound , thus inhibiting the growth of aerobic microorganisms                                                                                                                          

Phenols play an important biochemical role in prevention of infections by preventing the oxidation of arachidonic acid on the surface of the human cell and thus prevent the formation of mediators of inflammation as well as the ability of phenols to prevent oxidation processes caused by free radicals because the phenols reduce the free radicals to be inhibited and prevent the destruction of molecules rich in protein and nucleic acids.

 

B-    Detailed illustration:

Bio-bandages production starts by collecting shrimp shell wastes from Syrian fisheries and marine food restaurants (this is considered a positive recycling process for shell wastes). From these shells we extract a homo-polysaccharide called chitin. Shells are de-mineralized by treatweng wen acidic medium, then they are de-proteinated by moderate alkaline medium in order to remove proteins , calcium carbonate and calcium phosphate from the chitin and eventually, we obtain the chitosan after treating the chitin with highly concentrated alkaline medium which de-acetylises the chitin converting it into chitosan.

What is the chitosan ?

Chitosan is a fiber like substance derived from Chitin which is the second abundant polymer in the world after cellulose. Chitin and its de-acetylated derivative (chitosan) are natural homo-polymers composed of randomly distributed ß-(1→4) linked D-glucosamine (de –acetylated unit) and N-acetyl-D-glucosamine (acetylated unit). Chitosan is cationic bio-polymer and easily obtained from the exoskeleton of insects, marine creatures like shrimp, crab, lobster shells and fungal biomass.

Marine crustaceans consist of 30 – 40 % protein, 30 – 50 % calcium carbonate and calcium phosphate and 20 – 30 % chitin which is de-acetylated to get the chitosan.

Chitosan have got many biological properties such as biodegradation caused by enzymes released by such 7 % of soil – borne bacteria, in addition to the biocompatibility to human body cells. Chitosan is non-toxic, it was reported that Its oral LD50 (median lethal dose) in mice was found to be in excess of 16 g/day/kg body weight which is higher than that of sucrose.

The importance of chitosan  in wound healing : 

Chitosan degrades under the influence of enzymes present in body fluids such as lysozyme and N-acetylglucosoamidase. The degradation products, being chito-oligomers, are able to stimulate macrophages and positively influence collagen sedimentation, thus accelerating the wound healing process. A properly-selected chitosan also influences blood coagulability, and is therefore used in haemostatic dressings.

Efficacy of chitosan in the promotion of wound healing because chitosan has a positive charge; it attracts red blood cells, which have a negative charge. The red blood cells create a seal over the wound as they are drawn into the bandage, forming a very tight, coherent clot. In addition to providing hemostasis, chitosan polymer dressings also offer an antibacterial barrier.

One of the most important properties of Chitosan that enhance the possibility of chitosan usage in bio-dressings that analgesic effects for pain, the tested doses of different concentrations of a Chitosan solutions in acetic acid observed to reduce reactions associated with pain in mice, including the behavior of extended lists the background of the mouse and sclerosis abdominal that sounds to reflect the levels of pain.

 

                    Anit-bacterial and Anti-fungal activity of phenols:

The Anit-bacterial and Anti-fungal activity of olive pomace extracted phenols against three pathogenic fungi which are:

               ((Alternaria solani, Botrytis cinerea  and Fusarium culmorum

 As a result of a reported study, phenolic compounds extracted from olive leaves proved that they are promising natural killers for fungi and for various pathogens of agricultural crops.

In other study, it was reported that phenolic compounds extracted from olive leaves were tested against a number of bacteria and fungi that are considered infectious factors for digestive tract and respiratory tract in humans and they are the following:

 

µ  Positive gram bacteria:

(Bacillus cereus, B. subtilis and  Staphylococcus aureus)

µ  negative gram bacteria:

 (Pseudomonas aeruginosa , Escherichia coli and Klebsiella pneumonia)  

µ  fungi:

  (Candida albicans and Cryptococcus neoformans)

Seven phenolic compounds extracted from olive  were tested and they are namely :

caffeic acid, verbascoside, oleuropein, luteolin 7-O-glucoside, rutin, apigenin 7-O-glucoside and luteolin 4'-O-glucoside.

These phenolic compounds have shown individually and in combination high extraordinary effectiveness in growth inhibition of the tested strains which nominates the phenols extracted from the leaves of olive to be drugs can be used as antibiotics.

Chitosan – quinones dressings:

 Chitosan is naturally anti-microbial and the structure Chitosan – Quinones will significantly increase the anti – bacterial & anti-fungal properties of the Chitosan because it was proven that the phenolic compounds also possess anti-microbial activity and this ensures the ability of modified chitosan dressings to eliminate a wider range of pathogenic microorganisms that attack the wounds and burns as well as phenolic compounds that have antioxidant properties, the properties of anti-mutagenesis of cells, cancer-fighting properties and properties to control the rate of glucose in the blood.

Phenols play a biochemical role in prevention of the path of the interaction of inflammation in the body, which begins with the oxidation of (Arachidonic acid) - located in the wall of the human cells - by the (Lipoxyginase) enzyme that attacks the carbon-fifth of Arachidonic  acid in presence of oxygen yielding inflammatory mediators which are (Leukotrienes) compounds that bind to their receptors on the cell surface leading to release other inflammatory mediators such as histamine and thus increasing the permeability of vascular and pooling of blood in this region and this is one of the inflammation signs , the presence of phenols which are episodes of benzene linked to hydroxyl groups  reduce the oxygen and prevent the Lipoxyginase enzyme from oxidizing  of arachidonic acid . Therefore, phenols contribute to the prevention of inflammation.

 

 

 

                         Chitosan Applications

 

Applications of chitosan in the medicine:

µ Acceleration of wound healing without scar formation , haemostatic , pain blocking , tissue grafting and regeneration and can be used in deep & surface wounds

µ Gene delivery in gene therapy instead of bacterial plasmids

µ Anti-diabetic: lowering glucose levels and stimulating insulin secretion

µ Anti-tumor: causes cancer cells apoptosis

µ Immune system activator: stimulating resistance against systematic infections

µ Delivery of pharmaceuticals , vaccines , nutritive substances for patients in addition to developing their absorbance in the body

µ Chitosan films were suggested for kidney failure patients to act as artificial kidney membranes instead of natural kidneys because they possess an excellent permeability and tensile & compressibility tolerance

µ Tissue engineering: formation of three dimensional matrix in bone surgery to help in bone growth

µ Dentistry: formation of matrixes in tooth transplantation and bandages in tooth root surgery

µ Ophthalmology: production of colored and transparent contact lens

µ Burns treatment: chitosan films are permeable to oxygen in order to burn aeration

                       Applications of chitosan in the Agriculture:

1. Revitalizing agent for plant and soil: prevention of microbial infection and stimulating plant growth

2. Preservative packaging , chitosan is a bio-pesticide for nematode , bacteria , fungi and pathogenic insects , it is sprayed on seeds to elongate storing life

                 

 

 

 

 

                       Applications of chitosan in the food industry:

ü Edible and biodegradable films that are anti-bacterial and anti-fungal for food pathogenic factors because its ability to :

µ Controls moisture transfer between food and surrounding environment

µ Controls release of antimicrobial substances , antioxidants , nutrients, flavors and drugs

µ Reduction of oxygen partial pressure

µ Controls rate of respiration temperature control for food content

ü Food Additive material :

µ Clarification and de-acidification of fruits and beverages

µ Natural flavor extender

µ Emulsifying and thickening Color stabilization agent

Applications of chitosan in the cosmetic industry:

Chitosan is involved in substances used in cosmetic industries such as anti-sun creams because the chitosan absorbs ultra violet radiation victors from the sun and is a skin refreshing agent , preserves the hair color strength and stability and it can be used in tooth paste and mouth wash products.

Other applications of chitosan:

µ Heavy metal ions removal from polluted water

µ Paper industry: being it biodegradable , chitosan is used in packaging & wrapping paper and strengthens the recycled paper

µ Textile industries: chitosan removes undesired dyes

µ Chitosan is a chromatographic separating agent due to its chemical structure composed of chemical and functional groups that detect phenols and chloro-phenols……etc