Ulmo Honey & Vitamin C Treat Burn Wounds. Medicinal Honey Treatment

Medical Research Shows Ulmo Honey has a higher antibacterial effect compared to Manuka Honey.

"The choice of ulmo honey (Eucryphia cordifolia) for this research was based on its proven antibacterial properties. Sherlock et al. (2010) tested the antimicrobial activity of Chilean honey originating from ulmo against a group of bacteria, showing that this honey has a higher antibacterial effect for Staphyloccus aureus, Escherichia coli and Pseudomonas aeroginosa compared to European Manuka honey." Excerpt from the Discussion section for this study - see below. 

Comparative Study of Healing in Burns Treatment Based on Ulmo Honey (Eucryphia cordifolia) and Oral Vitamin C in Guinea Pig (Cavia porcellus) versus Hydrogel

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Contributors:
Carolina Schencke - PhD in Morphological Sciences, Universidad de la Frontera, Temuco, Chile.
Jessica Salvo - La Frontera, Temuco, Chile. - Faculty of Medicine, Universidad Mayor, Temuco Headquarters.
Adriana Vasconcellos - Faculty of Medicine, University of La Frontera, Temuco, Chile.
Mariano del Sol - Center for Research in Biomedical Sciences, Universidad Autónoma de Chile, Chile.

Information supplied by:
Universidad de La Frontera
Temuco - CHILE
Study Date: May 22 2013

Report listed in the International Journal of Morphology 
Original website permalink (spanish):
http://ref.scielo.org/5qhh5v
http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-95022013000300010&lng=en

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ABSTRACT:
A burn is a traumatic injury resulting in local and systemic injury with oxidative changes. Honey has antioxidant, anti-inflammatory and antibacterial properties. At the same time, among other benefits vitamin C improves the epidermal barrier and reduces wound contraction. The aim of this study was to evaluate morphologically the effect of regimen with Ulmo honey + vitamin C in guinea pigs and compare its healing and debriding potential with Hydrogel + tull in injuries caused by type B burns at day 10 post-injury. We used 15 guinea pigs (Cavia porcellus) distributed in groups A (Ulmo honey and vitamin C topical, oral), B (hydrogel-tull), and control (0.9% saline). Wound debridement was observed daily and wound treatment was performed daily until biopsies were obtained at day 10 post-injury. Four 4 um sections were stained with HE, Masson's trichrome and PAS. In group A there was rapid debridement when compared with groups B and Control. Treatment study presented advanced proliferative phase at day 10 post-injury with vascularized tissue, and presence of collagen fibers and fibroblasts. The epidermis was completely regenerated, presenting a visible basal lamina by PAS staining. Group B presented an initial proliferative fibroblastic phase, showing acute dermal elements. Epithelialization phase was completed in only 50% of the samples. Ulmo honey + vitamin C substances have been shown to be effective as treatment material in healing of type B burns in guinea pig skin.

INTRODUCTION

Burn is a traumatic wound resulting in both local and systemic damage, with obvious oxidative changes manifested by increased free radical activity. This type of injury oxidizes the tissue, increasing the enzyme xanthine oxidase and lipid peroxidation products. To these reactions are added free radicals produced by neutrophils (Gonzalez-Quevedo et al., 2001). Jureta, (2003), indicates that, in addition, the values ​​of superoxide dismutase, catalase, glutathione, alpha tocopherol and ascorbic acid decrease. Antioxidants required for cellular protection should be present in the daily diet, however, many of them are not available. In the study by Jawad et al. (2008), the oral administration of antioxidants in burn patients produced a significant improvement in the patients studied compared to the group of patients not treated with antioxidants. His study showed clearly that the use of oral antioxidants decreased the oxidative stress caused by burns.

Diaz (2003), in his Thesis on determining the floral origin of honeys labeled "honey of ulmo" (Eucryphia cordifolia), explains that knowing the botanical and geographic origin of honeys allows to establish a denomination of origin and give added value, Since many attributes and characteristics of the honeys depend on the vegetal species that the bees use as source of nectar for their elaboration. The uses of ulmo honey as a bactericide and fungicide have already been patented by Montenegro & Ortega, 2011, patent N WO / 2011/057421.

This is specified for the method of obtaining a phenolic extract of ulmo unifloral honey at 60% to be used as disinfectant and sanitizer.

In addition to having a large antimicrobial activity against various bacteria and fungi, the honey has an excellent antioxidant capacity, activating the monocytic line and anti-inflammatory factors (Bang et al., 2003; Buratti et al., 2007). Evans & Flavin, 2008). Studies in honey show histologically its anti-inflammatory activity (Van den Berg et al., 2008), demonstrating that it can be as effective as prednisolone (Bilsel et al., 2002). These authors describe that honey decreases the oxidative stress generated by the release of free radicals from burn injuries.

Stress associated with injury results in a greater need for Vitamin C. Stotts et al. (1990) analyzed a group of 90 individuals of average age 65, with second-degree injuries, where the vast majority showed a decrease in plasma vitamin and mineral values, mainly vitamin C. This vitamin demonstrates a variety of functions, among Which regulates the viability of the keratinocytes, epidermal barrier and basement membrane in vitro, reduces wound contraction after tissue implantation (Steven et al., 202). Rasik & Shucla (2000) demonstrated that adequate levels of antioxidants such as catalase, ascorbic acid and vitamin E play a crucial role in wound healing, especially in diabetic rats, or with liver and pancreatic and immunodepressed lesions.

The literature demonstrates beneficial healing, anti-inflammatory and antioxidant effects of the topical use of honey and Vitamin C in wound healing. However, there are few studies on its effect when both are associated. Subrahmanyam (1996) added antioxidants (vitamin C and E) plus polyethylene glycol (PEG) to honey, treating 42 patients partially burned. Patients treated with added honey were healed early in relation to patients treated with honey alone.

Depending on the extent of the wound, it will be necessary to use surgical treatments and frequent cures, for which there are multiple advanced dressings that promote the physiological moist environment necessary to achieve effective cicatrization that stimulates cell proliferation, cell migration Epithelial, debridement, bacterial and fluid control (Baranoski, 2008). Hydrogel is an autolytic debridant composed of thickening and wetting polymers plus water and absorbents (Andrades et al., 2004).

Advanced dressings are highly effective but cost more. Before this it is necessary to analyze other alternatives, within which is the honey of Chilean ulmo (Eucryphia cordifolia) with antioxidant and antibacterial properties.

This study aimed to morphologically evaluate the healing effect of a treatment based on honey plus ulna plus oral vitamin C in cuye (Cavia porcellus) and compared it with more synthetic tull hydrogel in skin wounds caused by type B burns.

This treatment alternative aims to be effective, fast and economical, which will be observed histologically in the regeneration of the wounds.

MATERIAL AND METHOD

Fifteen guinea pigs (Cavia porcellus) adults, 450 g of average weight, were kept with pellet ad libitum at the Experimental Surgery Center of the University of La Frontera, Temuco, Chile. The animals were divided into groups of 5, divided into groups A, treated with ulmochromic plus oral vitamin C, group B, treated with hydrogel and synthetic tull, and control group treated with 0.9% saline solution. The lesions, 0.7 cm in diameter, were induced on the back of the animals by contact with dry heat for 3 seconds, under intraperitoneal anesthetic effects with Atropine 40mg / kg, Ketamine 5mg / kg and Xylazine 0.05mg / kg. These covered deep skin, epidermis, hypodermis and part of the muscle tissue. Surgical edges were sutured and cured with serum until recovery.

The cures were performed applying warm physiological serum by syringe therapy 10 cm away from the lesion, and placing a gauze impregnated with Ulmo Honey in group A; Hydrogel plus tull in group B; And 0.9% saline solution in the control group. With this procedure the wounds were treated daily until the biopsies were obtained at the day 10 post-injury.

The biopsies were washed in saline, deposited in 10% buffered formalin, dehydrated and included in paraplast (Histosec-Merck®). Serial cuts of 10 mm were made until the injury and area cut in 4 mm thickness. For the histological analysis the sections were stained with HE and Masson trichomic (TCM). For histochemical analysis, we used periodic acid from Schiff (PAS). The slides were observed on an Olympus® model CX31 optical microscope, with camera model Moticam® Model 480. The images were projected onto a Sony® flat panel monitor. The percentage of polymorphonuclear cells (PMN), macrophages, active fibroblasts and inactive fibroblasts at day 10 post-injury was also determined in a total count of 100 cells in the scar area. The present study is part of the project DIUFRO N DI13-0044, and was accepted by the Ethics Committee of the University of La Frontera, Temuco, Chile.

RESULTS

The biopsy of healthy skin had thin epidermis, with thick corneal layer. The papillary dermis, superficial, is constituted by loose connective tissue, while in the deep, reticular dermis, it is constituted by abundant bundles of collagen distributed at random where you can see hair follicles and sebaceous glands. The biopsy of the test animal, burned on the back for 3 seconds, showed a deep damage of the skin, from the epidermis to the hypodermis. The biopsy of the control group, treated with physiological serum only, did not regenerate the epidermal layer at day 10 post-injury. Granulation tissue and edema were observed, presenting an acute inflammatory phase.

Group A (treated with honey plus oral vitamin C). At the moment of the injury, an immediate necrotic eschar by burn was observed, which causes spontaneous detachment on day 6 post-injury, leaving a wound of 0.4 ± 1 cm in diameter, not bleeding. Its diameter was reduced by about 50% a day 10 post-injury.

The biopsy showed an advanced proliferative phase at day 10 post-injury. The epidermis regenerated almost completely at day 10 post-injury, without yet presenting stratum corneum. The rest of the layers are differentiated: the well-developed basal stratum, differentiated spiny stratum and granular stratum outline. A basal lamina visible through PAS and TCM staining is observed. At the dermal level, a fibroblastic reaction was observed with abundant proliferation of collagen fibers, and neoformation of blood vessels. In this layer we observed scarcity or absence of hair follicles and sebaceous glands.

Biopsies obtained at day 10 of treatment with honey + vitamin C (A) and hydrogel (B), HE staining. A. The epidermis was regenerated with presence of basal, spiny stratum and granular stratum outline. B. The presence of the eschar in the lumen The dermis presented granulation tissue with abundant neoformation of blood vessels and extensive fibroblastic reaction with formation of collagen fibers.

Biopsies obtained at day 10 of treatment with honey + vitamin C (A) and hydrogel (B), Masson's trichrome stain. A. The regenerating dermis presented proliferation of collagen fibers and neoformation of blood vessels. The epidermis was regenerated observing the presence of basal lamina. B. The dermis presented abundant connective tissue in collagen fibers regularly arranged and neoformation of blood vessels. The epidermis was in development, with no basal lamina present.

Group B (treated with hydrogel + tull). The eschar generated by the burn detached spontaneously between 8-9 post-injury, leaving a bloody wound of granulatory appearance, bleeding on contact, without macroscopic evidence of epithelization. It had a diameter of 0.6 _ + 1 cm with a reduction of 15% at day 10 of the biopsy.

The biopsy extracted from the area treated with advanced dressings presented an initial proliferative fibroblast stage and acute inflammatory elements were still present. At the level of the dermal layer granulation tissue was observed, with intense fibroblastic reaction. The epidermis was not able to regenerate completely at day 10 post injury. If there was epidermal regeneration at the edges of the wound. The dermis presented an abundant connective tissue in collagen fibers regularly arranged and neoformation of blood vessels. The epidermis was differentiated, without the presence of basal lamina.

The percentage of active and inactive fibroblasts in the biopsies obtained at day 10 of treatment with honey ulmo + vitamin C and hydrogel. A percentage with 16% more active fibroblasts was observed in biopsies treated with hydrogel, which presented an initial proliferative phase of cicatrization.

DISCUSSION

Treatment with ulmo honey and oral vitamin C shortens the inflammatory phase in the regeneration process. The scarring reaction at 10 days post-injury is no longer acute, the PMNs were decreased, as well as the number of macrophages, there was presence of active fibroblasts being in a fairly advanced proliferative phase. The proposed treatment stimulates the formation of granular tissue and tissue proliferation adjacent to the lesion without showing signs of infection. In this way a quick and good quality healing was achieved.

Wound treatments should achieve good debridement to allow the formation of granular tissue and consequent epithelialization. Since the eschar with hydrogel was maintained until day 9 post-injury, the epidermal reparative process was slowed, epidermal closure being observed only in 50% of the biopsies. The biopsies that showed epidermal closure presented thin, still lacking the formation of the stratum corneum. Ulmo honey associated with oral vitamin C showed excellent results in autolytic debridement, as the eschar spontaneously evolved 3 days before the conventional treatment, resulting in an injury smaller than 0.1 cm in diameter. Rapid debridement accelerated epidermal growth, achieving maturity in a shorter period. The action debridante of the honey could be due to the activation of proteases in the tissue through the hydrogen peroxide. The metalloproteases can be activated by oxidation processes along with the inhibition of serines thus helping debridement of the wounds (Molan, 2004; Cook, 2008).

If the qualities of honey, such as antioxidant capacity (Buratti et al., Evans & Flavin and Bang et al.), Monocytic line activation, cytokine release, tumor factor alpha and interleukins (Tonks et al. , We add the benefits of vitamin C, with a high anti-inflammatory power, direct stimulation of the synthesis of collagen fibers, increased proliferation of fibroblasts in 4 times (Burke, 2004; Boyce et al., 2002), could be inferred That the mixed treatment of honey associated with vitamin C has a double benefit. His contribution to cicatrization of guinea-pig burn (Cavia porcellus) was to achieve a correct morphogenesis of the epidermal barrier, proliferation of fibroblasts, stimulation of basement membrane development, blood vessel neoformation and keratinocyte synthesis. In this way it was possible to quickly promote the closure of the wound caused by the burn.

The choice of ulmo honey (Eucryphia cordifolia) for this research was based on its proven antibacterial properties. Sherlock et al. (2010) tested the antimicrobial activity of Chilean honey originating from ulmo against a group of bacteria, showing that this honey has a higher antibacterial effect for Staphyloccus aureus, Escherichia coli and Pseudomonas aeroginosa compared to European honey Manuka.

The literature demonstrates beneficial effects of topical use of honey and Vitamin C on wound healing. However, there are few studies on their effect when they are associated. Subrahmanyam, 1996, added antioxidants (vitamin C and E) plus polyethylene glycol (PEG) to honey, treating 42 patients burned in partial depth. Patients treated with added honey were healed early in relation to patients treated with honey alone. Schencke et al., (2011), compared morphologically the effect of treatment of vitamin C-associated ulmo honey versus the use of honey alone, as an alternative healing in guinea-pig burns (Cavia porcellus). Their results showed that granular tissue formation, fibroblast activation and reepithelialization are faster in the vitamin C-treated honey group. Coinciding with these authors, our results presented a more advanced regenerative stage with the study treatment compared to Conventional treatment. Lesions treated with honey and vitamin C were found in advanced proliferative stage at day 10 post-injury. While with conventional treatment, healing was at an early proliferative stage, with a high concentration of active fibroblasts and blood vessels.

The guinea pig (Cavia porcellus) is a good experimental model to test the cicatrizant action of medicaments, since this one has less variability in skin thickness (González, 2002). In addition, this animal along with other primates and humans, do not generate or store their own vitamin C, as do other mammals.

Honey is a product of high nutritional value, (Ureña et al., 2007) however, it does not exist in Chile as a product of clinical application for topical use in wounds. In this study, honey associated with vitamin C achieves similarities and even improvements in its results when compared to treatment based on advanced dressings.

The results described can generate a research line in the treatment of wounds and burns, honey and vitamin C being ideal substances as a treatment material, easy to apply and remove.

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Boyce, S .; Supp, A .; Swope, V. & Warden, D. Vitamin C regulates keratinocyte viability, epidermal barrier, and basement membrane in vitro, and reduces wound contraction after grafting of cultured skin substitutes. J. Invest. Dermatol., 118: 565-72, 2002.

Burke, K. Photodamage of the skin: protection and reversal with topical antioxidants. J. Cosmet. Dermatol., 3: 149-55, 2004.

Buratti, S .; Benedetti, S. & Cosio. Evaluation of the antioxidant power of honey, propolis and royal jelly by amperometric flow injection analysis. Talanta, 71: 1387-92, 2007.

Díaz, C. Determination of floral origin and physical and chemical characterization of honey bee (Apis mellifera L.), labeled as "honey of ulmo" (Eucriphya cordifolia Cav.) Thesis presented to qualify for the degree of Lineciado in Agronomy. Faculty of Agrarian Sciences. Universidad Austral de Chile, 2003.

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González, E. Experimental models for the evaluation of the healing action of drugs. Rev. Cubana farm. 36 (3): 189-96, 2002.


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Schencke, C .; Except, J .; Veuthey, C .; Hidalgo, A. & del Sol, M. Healing in burns type AB-B using ulmo honey associated with oral vitamin C. Int. J. Morphol., 29 (1): 69-75, 2011.

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Information supplied by:
Universidad de La Frontera
Temuco - CHILE
Study Date: May 22 2013

Report listed in the International Journal of Morphology 
Original website permalink (spanish):
http://ref.scielo.org/5qhh5v

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