What’s Known

Natural compounds such as oleo-gum-resins have been traditionally applied to treat burn wounds. The topical application of a cream containing Boswellia on excision wounds affected different phases of wound healing, including collagen synthesis and wound contraction, leading to a faster healing process.

What’s New

In the treatment of burn wounds, the healing effect of a 40% Boswellia carteri formulation containing 0.72 mg of β-boswellic acid per 100 g was compared to silver sulfadiazine. On each day of the study, both healing time and healing indices were statistically comparable between the groups.

Introduction

Acute thermal injuries requiring medical care and surgical treatment are prevailing and burdensome. Burns cause the loss of around 18 million disability-adjusted life years and more than 250,000 fatalities annually. ¹ The prevalence is higher in low-income and developing countries. Children, women, and the elderly are high-risk groups, followed by people with comorbid conditions such as epilepsy, blindness, deafness, and diabetes. Patients with mental and psychiatric disorders or drug abusers are more likely to have self-inflicted burns. Large families, single parents, illiteracy, and unemployment are familial and social risk factors. ^{2 , 3} Unprivileged victims face increased complications and risks of permanent damage due to a lack of access to appropriate management, such as split-thickness skin grafting, fluid and electrolyte repletion, and infection control. ⁴

Traditional medical practices offer a wealth of information regarding the prevention, diagnosis, and treatment of illnesses. These holistic therapy modalities, which focus on the overall health of the entire body, are practiced broadly worldwide. ⁵ Due to their indisputable importance to public healthcare, the World Health Organization (WHO) encourages evidence-based research on the safety, efficacy, and quality of traditional medicines. ⁶ Moreover, natural products are considered respected choices in drug discovery for research on challenging diseases, such as cancer, pain, and acquired immunodeficiency syndrome (AIDS). ⁷ Burn injuries have historically caused severe damage to humans due to the inevitable exposure to intense sunlight and fire. Therefore, natural products have long been used to treat burn wounds. ⁸ The availability of medicinal plants in each region is a key factor in applying them for wound healing. For instance, Aloe vera gel, Carica papaya products, Lawsonia inermis formulations, and coconut oil are a few African folk remedies for burns. ⁹ Plants that promote the natural skin healing process can be helpful in wound treatment. Because of the complexity of the process, numerous mechanisms are involved during healing. For instance, the flowers of Achillea have anti-inflammatory and antibacterial effects. Camomilla recutita, as a mild astringent, promotes epithelialization and stimulates granulation tissue formation. Jasminum auriculatum leaves boost the tensile strength during the early phases of healing and accelerate mucopolysaccharide accumulation. Rosmarinus officinalis promotes granulation tissue regeneration, angiogenesis, and collagen deposition. ¹⁰

Numerous studies indicated that plant exudates such as gum-resins and oleo-gum-resins can heal wounds. Picea abies exudate also known as coniferous tree exudate was used to treat complicated surgical wounds. ¹¹ Pistacia atlantica resin oil and extracts were shown to considerably reduce wound size. ¹² The very valuable oleo-gum-resin of Boswellia (family Burseraceae), frankincense or olibanum, is obtained from the bark of the trees during dry seasons and contains 3-9% volatile oil, 60-70% resin, and 27-35% gum. ¹³ Boswellia oleo-gum-resin, which has antimicrobial, antioxidant, anti-inflammatory, and analgesic properties, is widely used for skin disorders, rheumatism, and central nervous system diseases. The topical application of a single-dose cream containing Boswellia on excision wounds affects various phases of wound healing, including collagen synthesis and wound contraction. Additionally, this product boosts the tensile strength of the wound, leading to a faster healing process. ¹⁴ Traditional Iranian Medicine (TIM) manuscripts mentioned Boswellia oleo-gum-resin as a burn wound healing agent. ¹⁵ In this study, one traditionally-originated and pharmaceutically-evaluated formulation from Boswellia carteri (B. carteri) was developed. Then, in a randomized clinical trial, the effectiveness of the sterilized B. carteri formulation (BCF) versus silver sulfadiazine (SSD) cream 1% in the treatment of second-degree burns was compared.

Material and Methods

Study Design and Patients

This double-blind, randomized clinical trial was conducted on 54 patients with second-degree burn wounds at Shiraz Burn Hospital, (Shiraz, Iran), from July 2012 to August 2013. Patients of both sexes aged 20 to 60 years old who had second-degree burn wounds of thermal origin affecting up to 5% of total body surface area (TBSA) were invited to take part in the study. Patients with known malignancy, collagen vascular disease, peripheral vascular disease, immunosuppressive disorders, cardiovascular diseases, diabetes mellitus, G6PD deficiency, electrical and respiratory burns, known allergy to the agents used, pregnancy, or breastfeeding were excluded. A further exclusion criterion was taking any medications that interfere with the wound healing process or affect the healing pace such as glucocorticoids, non-steroidal anti-inflammatory drugs, and chemotherapeutic drugs.

Eligible participants were randomly allocated into two groups. One group received SSD cream 1% produced by Sobhan Pharmaceutical Company (Iran), while the other received BCF 40%. The sample size for each group was determined using the means-comparison formula, with the mean healing time in the SSD group being 10±0.9 days. Using the below formula, and also considering α=0.05 and β=0.90, a sample size of 15 was calculated for each group. ¹⁶ Blocked randomization with a block size of two was applied. Time until complete healing was considered the primary outcome in the statistical analysis.

$n_1=n_2=\frac{(S_1^2+S_2^2){(Z_{1-\frac{\alpha }{2}}+Z_{\mathrm{1-\beta }})}^2}{{({\mu }_1-{\mu }_2)}^2}$

The study was approved by the Ethics Committee of Shiraz University of Medical Sciences (CT-90-5981) and also was registered in the Iranian Registry of Clinical Trials (IRCT201208041605N14). The participants were informed about the goals of the research as well as the whole process of the study. Written informed consent was obtained from the patients before participation. The study was carried out in agreement with the principles of the Helsinki Declaration and Iranian research ethical codes.

Preparation of Sterilized B. carteri Formulation

The oleo-gum-resin of B. carteri Birdw. was purchased from a local market in Shiraz (Iran). The samples were authenticated and deposited in the herbarium of the Phytopharmaceutical Department of the School of Pharmacy, Shiraz University of Medical Sciences (Shiraz, Iran) under voucher number PM400-5. This medicinal herb was chosen for research based on its market availability, safety, affordable price, and potential for a novel formulation that has not yet been reported. The sample was subjected to quality control tests. According to Food and Drug Administration regulations, any topical medication used to treat burns, wounds, and chronic ulcers must be sterile to avoid introducing exogenous microorganisms. ¹⁷ The oleo-gum-resin of B. carteri was frozen (-20 °C) for two hours before being powdered in an electric grinder and packaged in double-wrapped plastic bags for gamma sterilization. Based on the initial microbial count, radiation dosimetry was conducted. To prevent any interference with radiation-induced paramagnetic effects, packages were stored at room temperature in darkness for at least 72 hours after radiation (irradiator: 60 Co). We used gas chromatography-mass spectrometry to analyze the essential oils before and after irradiation and then compared the chromatograms to determine the effect of sterilization on the active components. ¹⁸ For aseptic preparation, the ingredients of the traditional medicine-based formulation (B. carteri oleo-gum-resin 40% in an oily duck fat base) were sterilized along with all devices and instruments using approved techniques and methods. ¹⁹

Standardization of B. carteri Formulation

High-performance liquid chromatography (HPLC) analysis of the formulation was performed by a Cecil (CE 4900) instrument (UK) equipped with an ultraviolet detector set at 210 nm and a C18 Eurospher column (250*4.6 mm, 5 μm) and a C18 Eurospher guard column (40*4.6 mm, 5 μm). ²⁰ The flow rate of the mobile phase (methanol) was 0.8 mL/min. The solvents were of analytical grade. The β-boswellic acid content of the formulation was quantified against standard commercial β-boswellic acid (Sigma-Aldrich, USA). We performed co-chromatography, compared the retention times of the compounds, and used the co-injection method to differentiate the peak of β-boswellic acid.

Burn Care and Assessments

After admission and primary preparation, wounds were washed with normal saline and dried using sterile gauze. Before providing any treatment, an expert emergency burn physician evaluated the general condition of the lesions. According to the related random code for each patient, wounds were covered either with SSD or BCF. Before application, the treatments were coded and packaged in identical containers, and each patient received individualized care.

The dressing was changed every two days until the wound was epithelialized completely. The same expert physician monitored the healing process for all patients. In the case of any drug reaction, the patient was removed from the trial and was given the required conventional care.

A nurse took photographs of the wounds every two days, and the files were coded. For further calibration of the images, a length scale was used. Both formulations were packaged similarly and coded to identify them. The margins of the wound were marked by a surgeon who was not informed of the patient’s treatment groups. After digitally scanning the tracings with Sigma Scan Pro 5 software (Systat Software Inc., California), the planimetry technique was used to calculate the wound area. ²¹ The wound healing index, defined as (A_wo- A_wn)/A_wo*100%, was used to calculate the healing rate, where A_wo is the initial wound area, and A_wn is the wound area on the n^th day. ²²

Statistical Analysis

Data were analyzed using SPSS 22.0 (SPSS, Chicago, IL), and the continuous data were expressed as Mean±SD, and categorical data were presented as frequency and percentage. Missing values were replaced via linear interpolation, which used the valid values before and after the missing value to interpolate. The amount of time until complete healing was considered the primary outcome and was evaluated by Kaplan-Meier survival analysis. The log-rank test was used to compare the survival times of the two groups. The independent samples t test was used to compare healing indices and the total number of healed patients between the two groups at each time interval. Cox proportional hazards regression was used to calculate the crude and adjusted hazard ratios (HRs) and 95% confidence intervals. In the analysis, age, sex, weight, cause of the burn, and TBSA were all considered potential confounders. Variables that changed the crude HR by more than 10% remained in the multivariable model. For all statistical analyses, P<0.05 was considered statistically significant.

Results

Standardization of B. carteri Formulation

According to the HPLC standardization procedure, each 100 g of the BCF 40% product contained 0.72 mg of β-boswellic acid (figure 1).

Figure 1. High-performance liquid chromatography (HPLC) chromatogram for the methanolic extract of β-boswellic acid (a); standard curve for β-boswellic acid (b); and HPLC chromatogram for the methanolic extract of Boswellia carteri formulation (c) is illustrated.

Patients’ Characteristics

Sixty-one patients were evaluated for eligibility, but seven of them did not meet the inclusion criteria. As a result, 54 patients with second-degree burn wounds participated in this randomized clinical trial (RCT). In total, 26 patients received SSD 1%, whereas 28 patients received BCF 40%. Due to changes in clinical circumstances or a desire to receive another type of treatment such as herbal formulations, eight patients from the SSD group and nine from the BCF group discontinued their treatment. One patient from the SSD group had a resistant infection that did not respond to SSD; thus, he was excluded. Four patients in the BCF group experienced hypersensitivity and had to quit the treatment process. Finally, 17 patients from the SSD group and 15 from the BCF group successfully completed the RCT. The CONSORT flow diagram of the study is shown in figure 2. The baseline characteristics of the patients are presented in table 1. The groups were similar in terms of age (SSD=34.70±13.36 years; BCF=36.20±13.49 years; P=0.75), weight (SSD=67.82±12.17 Kg; BCF=68.80±8.60 Kg; P=0.79), sex (P=0.46), cause of burn (P=0.69), and site of wounds (P=0.05). However, the TBSA burnt for the SSD group was 1.70±0.68%, which was significantly greater than that of the BCF group 1.20±0.41% (P=0.01).

Figure 2. This figure represents the CONSORT flow diagram of the study.

Wound Healing

In total, 17 patients in the SSD group and 15 patients in the BCF group were included in the final analysis. According to table 2, there was no significant difference between the mean (95% CI) healing times for the SSD group and the BCF group, which were 10.94 (9.03-12.85) days and 10.73 (9.23-12.23) days, respectively (P=0.71). As indicated in table 3, the Cox proportional hazards model revealed no significant difference between BCF and SSD in terms of the healing effect (adjusted hazard ratio=0.9 [95% CI:0.4-1.9]). The healing indices of the patients are compared across the two groups in table 4. The healing index of all patients in the BCF group reached 1 on day 17, but not in the SSD group until day 20. However, the P values revealed no significant differences in healing indices between the groups on any day. Figure 3 demonstrates the trend of healing indices for patients in the two groups. Another variable that was compared in the SSD and BCF groups, was the number of patients who had recovered (figure 4).

Figure 3. The trends of healing indices for patients in the silver sulfadiazine cream 1% group and Boswellia carteri formulation group were compared with each other. P values are mentioned in table 4.

Figure 4. The trends of healed patients (%) in the silver sulfadiazine cream 1% group and Boswellia carteri formulation group were compared with each other.

Discussion

In this study, the healing effect of a 40% BCF containing 0.72 mg of β-boswellic acid per 100 g was compared against SSD in the treatment of burn wounds. On each day of the trial, both healing time and healing indices were statistically similar between groups.

Herbal products are widely available and culturally accepted in many areas, although there is little scientific proof of their safety and effectiveness. Traditional Iranian Medicine (TIM) offers a variety of herbal remedies for wound healing. ⁸ These products have anti-bacterial, anti-inflammatory, and anti-oxidative properties, which reduce the amount of time needed for healing. ²³ Finding evidence to support the rational usage of herbal medicines is receiving more attention from researchers, but the field demands more quantitative evaluations such as evidence-based clinical trials. ¹⁰ The SSD, a common treatment for burn wounds, just prevents infections and has no definite effect on the wound healing process. Some studies reported that SSD delayed the wound healing process due to its toxic effects on fibroblasts and keratinocytes. ^{24 , 25} Some of its side effects included silver staining, allergic reactions, and methemoglobinemia, and it is not recommended during pregnancy or infancy. ^{26 , 27} Therefore, looking for an alternative medication that does not have the limitations of SSD seems helpful. The oleo-gum-resin of B. carteri consists of boswellic acids and essential oils, composed mainly of octyl acetate, incensole, and incensole acetate. ^{18 , 28} The anti-inflammatory effect of boswellic acids were established in many studies. ^{29 , 30} Furthermore, the essential oils of Boswellia species act as antimicrobial agents. ^{31 , 32} Therefore, it is hypothesized that the wound-healing properties of Boswellia formulations are mostly due to their anti-inflammatory components such as boswellic acids and essential oils.

To the best of our knowledge, this is the first clinical trial to evaluate the burn wound healing effect of a sterilized product formulated from B. carteri. In this study, a topical formulation based on manuscripts from traditional medicine was developed. The main ingredient of the formulation, B. carteri oleo-gum-resin, was sterilized using gamma-irradiation to minimize the hazard of pathogenic microorganisms.

In 2004, a case of allergic contact dermatitis was reported in which Boswellia oleo-gum-resin was considered a safe herbal medicine. ³³ Essential oils are proven to be a potential source of allergic reactions. ³⁴ As B. carteri contains essential oils, the reaction may be associated with these compounds. Nevertheless, the findings of the present study indicated that utilizing BCF 40% could lead to the possibility of allergic contact dermatitis. Furthermore, exposure to gamma radiation of B. carteri might cause degradation of some components to low molecular weight compounds that penetrate the epidermal barrier faster and trigger hypersensitive reactions. The only limitation of the study was the difference between the TBSA of the two groups (P=0.01).

Conclusion

The findings of the present study indicated that a 40% BCF had healing activity in treating second-degree burn wounds. According to the results of this randomized clinical trial, BCF and SSD, the standard treatment for burn wounds, provide comparable healing times and healing indices. However, the likelihood of allergic reactions to BCF should be taken into consideration. Consequently, further studies on formulations with a lower concentration of B. carteri are suggested as products with lower percentages of oleo-gum-resin might not elicit hypersensitivity reactions.

Acknowledgment

This research, derived from the PhD dissertation of P. Badr, was supported by Shiraz University of Medical Sciences (grant no. 5981). The authors gratefully acknowledge the assistance of the Clinical Research Development Center of Namazee Hospital. They would like to express their gratitude to all participants as well as the hospital personnel for their sincere cooperation with the researchers.

Authors’ Contribution

P.B: conception, study design, acquisition of data, drafting the manuscript; S.A: conception, revising the manuscript critically; H.R.T: analysis and interpretation of data, revising the manuscript critically; A.A.M: study design, revising the manuscript critically; S.D: conception, study design, revising the manuscript critically; All authors have read and approved the final manuscript and agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

Conflict of Interest

None declared.

References

1. Rybarczyk MM, Schafer JM, Elm CM, Sarvepalli S, Vaswani PA, Balhara KS, et al. A systematic review of burn injuries in low- and middle-income countries: Epidemiology in the WHO-defined African Region. Afr J Emerg Med. 2017; 7:30-7. Publisher Full Text | DOI | PubMed
2. Peck MD. Epidemiology of burns throughout the world. Part I: Distribution and risk factors. Burns. 2011; 37:1087-100. DOI | PubMed
3. Smolle C, Cambiaso-Daniel J, Forbes AA, Wurzer P, Hundeshagen G, Branski LK, et al. Recent trends in burn epidemiology worldwide: A systematic review. Burns. 2017; 43:249-57. Publisher Full Text | DOI | PubMed
4. James SL, Lucchesi LR, Bisignano C, Castle CD, Dingels ZV, Fox JT, et al. Epidemiology of injuries from fire, heat and hot substances: global, regional and national morbidity and mortality estimates from the Global Burden of Disease 2017 study. Inj Prev. 2020; 26:i36-i45. Publisher Full Text | DOI | PubMed
5. Lemonnier N, Zhou G-B, Prasher B, Mukerji M, Chen Z, Brahmachari SK, et al. Traditional knowledge-based medicine: a review of history, principles, and relevance in the present context of P4 systems medicine. Progress in preventive medicine. 2017; 2:e0011. DOI
6. Organization WH. WHO traditional medicine strategy: 2014-2023. World Health Organization: Geneva; 2013.
7. Taylor PW. Alternative natural sources for a new generation of antibacterial agents. Int J Antimicrob Agents. 2013; 42:195-201. DOI | PubMed
8. Badr P, Daneshamouz S, Mohammadi AA, Afsharypuor S. Knowledge of burn wound healing: the heritage from traditional pharmacy of Persia. Pharm Hist (Lond). 2014; 44:88-93. PubMed
9. Albertyn R, Berg A, Numanoglu A, Rode H. Traditional burn care in sub-Saharan Africa: a long history with wide acceptance. Burns. 2015; 41:203-11. DOI | PubMed
10. Maver T, Maver U, Stana Kleinschek K, Smrke DM, Kreft S. A review of herbal medicines in wound healing. Int J Dermatol. 2015; 54:740-51. DOI | PubMed
11. Ghodela NK, Prasad P, Kumar V, Dudhamal T. Wound healing potential of gums &amp; oleo-gum-resins: a brief review. Global J Res Med Plants & Indigen Med. 2017; 6:89-94.
12. Shahouzehi B, Sepehri G, Sadeghiyan S, Masoumi-Ardakani Y. Ameliorative effects of Pistacia atlantica resin oil on experimentally-induced skin burn in rat. Res J Pharmacogn. 2019; 6:29-34. DOI
13. Wichtl M. Teedrogen und phytopharmaka. Wissenschaftliche Verlagsgesellschaft: Stuttgart; 2009.
14. Bansal N, Mehan S, Kalra S, Khanna D. Boswellia serrata-frankincense (A Jesus Gifted Herb); an updated pharmacological profile. Pharmacologia. 2013; 4:457-63. DOI
15. Aghili Shirazi S. Makhzan al advieh. Rah-e Kamal: Tehran; 2011.
16. Campanati A, De Blasio S, Giuliano A, Ganzetti G, Giuliodori K, Pecora T, et al. Topical ozonated oil versus hyaluronic gel for the treatment of partial- to full-thickness second-degree burns: A prospective, comparative, single-blind, non-randomised, controlled clinical trial. Burns. 2013; 39:1178-83. DOI | PubMed
17. Eaglstein WH, Kirsner RS, Robson MC. Guidance for industry chronic cutaneous ulcer and burn wounds-developing products for treatment. Wound repair and regeneration. 2012; 20:793-6.
18. Badr P, Daneshamouz S, Mohammadi A, Akbarizadeh A, Afsharypuor S. The effect of 60Co-gamma radio-sterilization on Boswellia carterii essential oil composition. Research Journal of Pharmacognosy. 2016; 3:67-74.
19. Revision USPCCo. United States Pharmacopeia, the National Formulary. Chap. 31. sterility: Microbial tests; 2008.
20. Wang C, Xia L, Song Z, Li Q, Wang C, Zeng L, et al. [Determination of five boswellic acids in Boswellia serrata]. Zhongguo Zhong Yao Za Zhi. 2011; 36:1330-3. PubMed
21. Khoo R, Jansen S. The Evolving Field of Wound Measurement Techniques: A Literature Review. Wounds. 2016; 28:175-81. PubMed
22. Horn SD, Barrett RS, Fife CE, Thomson B. A predictive model for pressure ulcer outcome: the Wound Healing Index. Adv Skin Wound Care. 2015; 28:560-72. DOI | PubMed
23. Dan MM, Sarmah P, Vana DR, Dattatreya A. Wound healing: concepts and updates in herbal medicine. Int J Med Res Health Sci. 2018; 7:170-81.
24. Raymond SL, Zecevic A, Larson SD, Ruzic A, Islam S. Delayed Healing Associated with Silver Sulfadiazine Use for Partial Thickness Scald Burns in Children. Am Surg. 2018; 84:836-40. PubMed
25. Khansa I, Schoenbrunner AR, Kraft CT, Janis JE. Silver in Wound Care-Friend or Foe?: A Comprehensive Review. Plast Reconstr Surg Glob Open. 2019; 7:e2390. Publisher Full Text | DOI | PubMed
26. Marx DE, Barillo DJ. Silver in medicine: the basic science. Burns. 2014; 40:S9-S18. DOI | PubMed
27. Genuino GA, Baluyut-Angeles KV, Espiritu AP, Lapitan MC, Buckley BS. Topical petrolatum gel alone versus topical silver sulfadiazine with standard gauze dressings for the treatment of superficial partial thickness burns in adults: a randomized controlled trial. Burns. 2014; 40:1267-73. DOI | PubMed
28. Woolley CL, Suhail MM, Smith BL, Boren KE, Taylor LC, Schreuder MF, et al. Chemical differentiation of Boswellia sacra and Boswellia carterii essential oils by gas chromatography and chiral gas chromatography-mass spectrometry. J Chromatogr A. 2012; 1261:158-63. DOI | PubMed
29. Ammon HP. Boswellic Acids and Their Role in Chronic Inflammatory Diseases. Adv Exp Med Biol. 2016; 928:291-327. DOI | PubMed
30. Iram F, Khan SA, Husain A. Phytochemistry and potential therapeutic actions of Boswellic acids: A mini-review. Asian Pacific journal of tropical biomedicine. 2017; 7:513-23. DOI
31. Sadhasivam S, Palanivel S, Ghosh S. Synergistic antimicrobial activity of Boswellia serrata Roxb. ex Colebr. (Burseraceae) essential oil with various azoles against pathogens associated with skin, scalp and nail infections. Lett Appl Microbiol. 2016; 63:495-501. DOI | PubMed
32. Mohamed AA, Ali SI, Kabiel HF, Hegazy AK, Kord MA, EL-Baz FK. Assessment of antioxidant and antimicrobial activities of essential oil and extracts of Boswellia carteri resin. Int J Pharm Phytochem Res. 2015; 7:502-9.
33. Acebo E, Raton JA, Sautua S, Eizaguirre X, Trebol I, Perez JL. Allergic contact dermatitis from Boswellia serrata extract in a naturopathic cream. Contact Dermatitis. 2004; 51:91-2. DOI | PubMed
34. Reeder MJ. Allergic Contact Dermatitis to Fragrances. Dermatol Clin. 2020; 38:371-7. DOI | PubMed