Effects of Three Types of Honey on Cutaneous Wound Healing

Hatice Ozlem Nisbet, DVM, PhD; Cevat Nisbet, DVM, PhD; Murat Yarim, DVM, PhD; Ahmet Guler, PhD; Ahmet Ozak, DVM, PhD



In This Article


Ethical Approval

The study was carried out in accordance with the Canadian Council on Animal Care (CCAC) Guidelines and the Declaration of Helsinki. The experimental protocol was approved by the Experimental Animal Studies Ethics Committee of Ondokuz Mayis University and also by the local ethics committee approved by the Central Experimental Animal Studies Ethics Committee of the Republic of Turkey.

Study Population

A total of 24, 6-month-old, New Zealand white female rabbits (2500 g ± 300 g body weight), which had been supplied by the Animal Unit at Ondokuz Mayis University, Samsun, Turkey, were used in the study. Upon arrival at the institution, the animals were housed in an environmentally controlled animal facility for 7 days for acclimatization. Each rabbit was housed in its own standard cage and subjected to 12 hours of light and 12 hours of darkness. The room temperature and humidity were maintained at 19°C ± 1°C and 55% ± 10%, respectively. All rabbits were fed 160 g pelleted rabbit diet daily and water was available ad libitum. Care was taken to avoid unnecessary stress and discomfort to the animals throughout the experiment. The Ondokuz Mayis University Animal Care and Use Committee approved the study protocol.

A complete blood cell count was performed for each rabbit on days 0, 7, 14, and 21.

The types of honey used in this study were pure chestnut (Castanea sativa) honey, pure rhododendron (Rhododendron luteum) honey, and pure blossom (multifloral) honey. The honeys were procured from the Apiary of Ondokuz Mayis University and from local beekeepers in the Black Sea Region of Turkey. Standard rearing methods were applied to the colonies.[12,13] In April, 7 kg–8 kg of syrup was given to each colony to ensure adequate nutrition. Beeswax cake, syrup, or chemicals to treat honeybee diseases were not given to the colonies during the main nectar flow period. The main honeycomb and supper were given to the colonies on an as needed basis. While chestnut and rhododendron honeys were produced in the Turkeli (41°N and 34'E) district of Sinop where these plants are common, the blossom honey was produced in the Kelkit district of Gumushane (39°N 29'E), which was rich in thyme (Satureja thymbra L), labiatae (Lamium album), alfalfa (Trifolium ambiguum), and geven (Astragalus microcehalus) blossoms. Honeys were produced in the months of August and September. The honeys were extracted and wrapped following filtration with a 0.2-mm filter.[12]

Wound Creation

Before the surgery was performed, the animals were premedicated using intramuscular (IM) xylazine (7 mg/kg, [Rompun®, Bayer, Istanbul, Turkey]) and anesthetized with IM ketamine (40 mg/kg, [Ketasol®, Richterpharma, Interhas, Ankara, Turkey]). A single dose of cephazolin sodium (30 mg/kg IM, [Cefozin® Bilim, Istanbul, Turkey]) was administered for antibiotic therapy preoperatively. Carprofen (4 mg/kg, subcutaneously [SC], [Rimadyl®, Pfizer Inc, Zaventem, Belgium]) was injected in all animals once just prior to the operation and every 24 hours for 3 days postoperatively.

Each rabbit was positioned in sternal recumbency. After clipping the hair on the back of the rabbits, the skin was sterilized with polyvidone-iodine (Betadine®, Kansuk, Istanbul, Turkey). The skin and the underlying cutaneous trunci muscle were excised with a #11 scalpel and scissors to create four 1.5 cm x 1.5 cm full-thickness wounds in 4-cm intervals. Hemorrhage was controlled by sterile surgical sponge compresses. One wound from each animal (test wound) was assigned randomly to receive PCH (0.5 mL), PRH (0.5 mL), or PBH (0.5 mL) with the fourth wound serving as the untreated control. After application of the topical treatments, the wound areas were then bandaged with sterile nonadherent pads and porous adhesive tape. The wounds were cleansed with sterile saline solution and the topical applications were done every other day until complete epithelialization was achieved. Eight rabbits were evaluated on days 7, 14, and 21, respectively. Four wounds were created on each rabbit for a total of 32 wounds. Wound data were obtained at each application time; therefore, n values were accepted as 32 for histopathological, biochemical, and planimetry analyses. Five-millimeter (5-mm) punch biopsy instruments were used to obtain two skin specimens from the wound edge of each rabbit, using planimetry, immediately after measurement was performed. The skin specimens were sent to the laboratory for pathological and biochemical analysis.

Subjective Observations

At every bandage change, the wounds were observed grossly for redness, swelling, odor, edema, vesiculation of the wound, marked accumulation of exudate, and scab formation. The wounds were also observed for formation of exuberant granulation tissue (more than the wound edges) during the study.

On days 14 and 21 any hair that had grown around the wounds was trimmed. The day that initial granulation tissue was observed, the day that the wound was covered, and the day the wound was completely filled with granulation tissue and epithelialized were recorded. The observations were performed in a nonblinded manner.