This section encompasses the literature published in the last years describing the application of naturally occurring antimicrobials, anti-inflammatory and regenerative molecules in the acceleration of the wound-healing process. A wealth of in vitro and in vivo studies with animal models demonstrates the antimicrobial, anti-inflammatory, and regenerative properties of essential or edible oils, honey, aloe vera, plant extracts, cationic peptides, etc.; however in the sake of concision we shall only review those results which involve clinical trials. Results from enteral administration of these natural origin components have also been excluded in this review.
4.1. Antimicrobials
Everything has antimicrobial properties depending on its dose. “The dose makes the poison” is the basic principle of toxicology (credited to Paracelsus). The idea behind this section is to be thought-provoking, trying to compare the potential benefits from natural-origin materials topically applied against pathogenic microorganisms colonizing wounds compared to the conventional synthetic topical antiseptics (iodine, silver, chlorhexidine, polyhexamethylbiguanide, etc.) or with the systemic application of antibiotics and also always keeping in mind the required dose needed to reach the same antimicrobial level. It is important to mention that to prevent wound infection the use of topical antibiotics is not recommended due to the risk of sensitization and development of resistance. Natural origin materials have been used in wound care with positive results.
Natural origin materials can be antimicrobials or can boost the defenses of the infected host. As we mentioned before, when the antimicrobial is applied on the wound or on intact skin they are called antiseptics [
136]. Obviously the material of choice to accelerate the physiological wound healing process must have antimicrobial action on bacteria, yeast, fungi, virus, and spores but should be non-cytotoxic on human cells without the development of any antimicrobial resistance.
Honey has been used as antimicrobial for centuries. The composition of the honey depends on the floral source that the honeybees use and also on the environmental conditions [
137]. Its antimicrobial action is attributed to its acidic pH, high osmolarity, and to the presence of hydrogen peroxide (reactive oxygen species (ROS) generation), antioxidants, lysozyme, polyphenols, phenolic acids, flavonoids, methylglyoxal, and bee peptides [
137]. A prospective, multicentre, open label randomized controlled trial with 808 patients with venous leg ulcers having ≥50% wound area covered in slough not taking antibiotics or immunosuppressant therapy were recruited to compare the efficacy of manuka honey versus current hydrogel therapy [
138]. The results showed that patients treated with the manuka honey had increased incidence of healing, effective desloughing and a lower incidence of infection than the controls. However, Jull
et al. [
139] carried out a community-based open-label randomized trial on patients with venous ulcers using either calcium alginate dressings impregnated with manuka honey (187 patients) or usual care (181 patients) and showed that honey-impregnated dressings did not significantly improve venous ulcer healing at 12 weeks compared with the usual care. Another prospective open label multicenter study with 108 patients with sloughy venous leg ulcers treated with manuka honey or hydrogel showed a benefit of one versus the other depending on the bacteria present [
140]. A randomized study with 69 patients using honey-coated bandages compared with silver-coated bandages on the treatment of malignant wound showed no differences between both groups [
141]. In a prospective randomized study (45 subjects) to compare the effectiveness of honey dressings vs. povidone iodine dressing in chronic wound healing, honey dressings showed a significant decrease in the wound surface area, pain score and increase in comfort score compared to the iodine-based ones [
142]. Shorter times of healing and a rapid disinfection of neuropathic diabetic foot ulcers were observed when using manuka honey-impregnated dressings compared to conventional dressings in a randomized clinical trial (63 patients) [
143]. Literature reports varied results for honey depending on the infected wound [
144] and also it has been demonstrated that honey may even have a detrimental effect on diabetic ulcers [
145]. These contradictory results indicate that more information is needed and multi-center clinical trials and with a larger number of patients are required to show a clinical benefit for honey to reduce the incidence of wound infection [
146].
Essential oils are composed of 20–80 constituents existing at significantly low concentrations in plants which chemical composition depends on climatic, seasonal, geographic conditions and distillation technique [
147]. Their antimicrobial action is attributed to some of their varied components including terpenes, low molecular weight aliphatic hydrocarbons, acids, alcohols, aldehydes, acyclic esters,
etc. Different edible oils extracted from fruits have antimicrobial action which is attributed to their acidic pH and to the presence of simple phenols and oxygenated compounds. The great advantage is that essential oils show little impact on the development of antimicrobial resistance and susceptibility compared to other biocidal components [
148,
149].
Darmstadt
et al. [
150] demonstrated that premature babies (159) who received a daily massage with sunflower-seed oil were 41% less likely to develop nosocomial infections than controls (without any treatment) and Aquaphor (petrolatum, mineral oil, mineral wax, lanolin alcohol; 157 babies) did not significantly reduce the risk of infection.
A randomized, controlled trial of tea tree topical preparations versus a standard topical regimen for the clearance of methicillin-resistant
S. aureus (MRSA) colonization was carried out by Dryden
et al. [
151]. In this study 114 patients received standard treatment and a 49% of them were cleared of MRSA carriage. 110 patients received tea-tree oil regimen and only a 41% of them were cleared. The authors concluded that there was no significant difference between treatment regimens and, from the same study they also concluded that tea-tree treatment was more effective than chlorhexidine or silver sulfadiazine at clearing superficial skin sites and skin lesions. A recent prospective, open-label, randomized, controlled trial with 445 patients was carried out to determine whether the daily use of 5% tea-tree oil body wash compared with standard care had a lower incidence of MRSA colonization [
152]. The results showed that a 10% of the patients developed new MRSA colonization and therefore, compared with standard care, the daily use of 5% tea-tree oil body wash cannot be recommended as an effective means of reducing MRSA colonization.
Essential coriander oil was very efficient as an antiseptic for the prevention and treatment of skin infections with Gram-positive bacteria (
Streptococcus pyogenes,
S. aureus and MRSA) [
153]. In addition, no skin irritation could be observed by sensitive photometric assessment in any of the 40 volunteers using patches impregnated with this essential oil.
Lavender oil was used in the treatment of recurrent aphthous ulceration by means of randomized double-blind, placebo-controlled study performed in animal models and also in 115 subjects [
154]. A significant ulcer-size reduction, increased rate of mucosal repair, and healing within three days of treatment were observed in the animals treated with lavender oil compared to baseline and placebo groups. Lavender oil showed a broad antibacterial activity against different tested strains and the patients treated with lavender oil showed a significant reduction in inflammation level, ulcer size, healing time, and pain relief. As in the case of honey-based products, the literature compiles contradictory or at least insufficient results to make a scientific conclusion about the demonstrated benefit of those essential oils.
Antimicrobial peptides are an evolutionarily conserved component of the innate immune response found among all classes of life ranging from prokaryotes to humans which show antimicrobial activities against Gram-positive and Gram-negative bacteria [
155]. They act with multiple roles as mediators of inflammation with the effects on epithelial and inflammatory cells, influencing cell proliferation, wound healing, cytokine/chemokine production and chemotaxis [
156]. They also do not seem to propagate the development of antibiotic-resistant micro-organisms [
157]. Lipsky
et al. [
158] described the results of two consecutive, double-blind, randomized controlled trials on diabetic patients with a mildly infected diabetic foot ulcers to receive an active topical antimicrobial peptide, pexiganan acetate cream, or an active oral antibiotic (ofloxacin), plus a respective inactive placebo. The results with a total of 835 subjects showed equivalent results (within the 95% confidence interval) for topical pexiganan and oral ofloxacin in clinical improvement rates, overall microbiological eradication rates, and wound healing rates. A significant reduction of infectious complication after major liver surgery was also observed in a clinical trial with patients using bactericidal/permeability-increasing protein (rBPI (21)) [
159].
Despite some of those good results, the main drawback is their high cost and also that it is difficult to maintain a constant optimal therapeutic level due to the short half-life of recombinant proteins
in vivo [
160].
Several pathogenic bacteria are developing antibiotic resistance and therefore, initially the study of the potential applicability of those natural occurring antimicrobial agents is justified [
161,
162]. However, as in the case of nanosilver, more studies are need to assess their potential role in antimicrobial resistance [
163].
4.3. Regenerative
Healing advances were observed in a clinical trial with 120 primiparous women, with singleton pregnancy, who were treated during episiotomy recovery either with lavender oil or with povidone-iodine (controls). The results concluded that there was no significant difference between those two groups in surgery-site complications. However, redness in lavender group was significantly less than in controls [
168]. Another clinical trial with 89 women was carried out to analyze the recovery after episiotomy using lavender based-on olive oil and olive oil [
169]. The results also suggested that lavender based-on olive oil and olive oil should be added to routine water sitz bath for post-episiotomy care. In another clinical trial involving 111 primiparous women, Eghdampour
et al. [
170] demonstrated that women treated either with aloe vera or with calendula ointment showed a faster episiotomy wound healing compared to the untreated group.
A clinical study involving 30 patients treated with sesame oil or just with saline having fresh traumatic wounds showed that this oil was effective by reducing pain, minimizing wound surface, reducing the discharge and promoting the epithelialization compared to the controls [
171].
Aloe vera gel has been traditionally used to treat burn wounds and several studies concluded that cumulative evidence tends to support that aloe vera might be an effective interventions used in burn wound healing for first to second degree burns but the same study states that more studies are needed to corroborate it [
172]. A systematic review of the literature including clinical trials concluded that controlled clinical trials in humans demonstrated no benefit when aloe vera was incorporated into topical therapy [
173].
A prospective randomized double-blind clinical trial was conducted with 90 women who had undergone cesarean operation applying aloe vera or just a simple dressing. A significant difference was observed between the two groups with respect to the wound healing score 24 h after the operation; however after eight days, the difference in the wound healing score was not significant [
174].
A prospective clinical trial was conducted with 60 patients to evaluate the effects of a topical cream containing 0.5% aloe vera juice powder in the treatment of chronic anal fissures by Rahmani
et al. [
175]. The study concluded that there were statistically significant differences in chronic anal fissure pain, hemorrhaging upon defection and wound healing before and at the end of the first week of treatment with the gel in comparison with the untreated group (placebo). A comparative study stablished between second degree burn patients (50) treated with aloe vera gel compared with those treated with 1% silver sulphadiazine cream showed that faster wound healing and less pain were reported for the group treated with the aloe [
176]. Once again we found controversial results or incomparable results because the effect of any antiseptic depends on the wound type, chronicity, the moment to apply the therapeutic compound, the condition of the patients, origin, age, combinatory effects, contact time, the antiseptic of choice for a particular (e.g., bacterial strain) infection,
etc.
Propolis, a resinous mixture that honey bees collect from tree buds, sap flows, or other botanical sources, was also used in a clinical trial to study its influence in the treatment of recurrent aphthous stomatitis, a common, painful, and ulcerative disorder of the oral cavity [
177]. The results obtained indicated that patients in the propolis group self-reported a significant improvement in their quality of life and a statistically significant reduction of outbreaks compared to the untreated controls.
A randomized clinical trial of 37 patients with neuropathic diabetic foot ulcers was set to analyze the benefits of extract of kiwifruit compared to a standard dressing treatment [
178]. The group treated with the kiwifruit experienced a larger reduction in the size of the ulcer than the untreated one at the same time. The amount of collagen, angiogenesis, vascularization and granulation tissues were significantly higher in the experimental groups than in the controls. Wound healing kinetics were also evaluated on 34 patients with chronic venous leg ulcers treated either with the plant species
Ageratina pichinchensis or with 7% propylene glycol alginate [
179]. The results showed that the plant extract produced a 100% therapeutic effectiveness, while the control treatment achieved this condition in 81.8% of the control group patients. Ulcer size reduction was significantly higher in the group of patients administered with the extract.
Banana leaf, tree bark, cocoa, turmeric, β-glucans,
etc. have also demonstrated their antioxidant, anti-inflammatory, antimicrobial, and angiogenic properties in
in vitro studies and, for some of them, in clinical trials [
180].
There is an endless list of natural origin products which have been used since antiquity to heal wounds, but still scientific evidence of those properties is still to be demonstrated using double-blinded multi-center randomized placebo-controlled trials to reach a sufficient level of evidence.
4.4. Dose Analysis
To compare the doses needed to obtain the same output in the prevention of infection during wound healing when using natural origin components or synthetic ones we compared the minimal inhibitory and minimal bactericidal concentrations (mic and mbc, respectively) published for just honey in the sake of concision. In this regard Tan
et al. [
181] calculated the mics for two different honeys against nine different bacteria (
S. pyogenes,
coagulase-negative Staphylococci,
MRSA,
Streptococcus agalactiae,
S. aureus ATCC 33591,
Proteus mirabilis,
Shigella flexneri,
Escherichia coli and
Enterobacter cloacae) and the values ranged from 8.75% (w/v) to 25% for tualang honey, while those for manuka honey ranged between 8.75% and 20% (w/v). The mics and mbcs reported by Henriques
et al. [
182] when challenging
S. aureus NCTC to manuka honey were 2.9% (w/v) and 4.5% (w/v) as mean mic and mbc values, respectively. Against
P. aeruginosa ATCC 27,853 the doses required of manuka honey were 9.5% (w/v) and 12% (w/v), mic and mbc respectively.
Mic and mbc doses of 20% (w/v) and 40% (w/v), respectively were required when using manuka honey against biofilm forming strains of
S. pyogenes MGAS6180 [
183]. Against oral bacteria (
Streptococcus mutans adhered on a glass substrate), Badet and Quero [
184] reported a total inhibition of multi-species biofilm at the concentration of 200 μg/mL using manuka honey and a total biofilm inhibition was reached at a concentration of 500 μg/mL. Propolis was much more effective against
S. aureus and
Escherichia coli compared with honey as reported by Rahman
et al. [
185]. These authors estimated that propolis at concentrations of 2.74 to 3.5 and 3.5 mg/mL is effective to inhibit
S. aureus and
E. coli, respectively. On the contrary, honey was effective to inhibit
S. aureus at the concentration of 375 mg/mL but failed to inhibit
E. coli growth at same concentration.
Different Malaysian honeys were evaluated against
S. aureus,
Bacillus cereus,
E. coli, and
P. aeruginosa and compared using equivalent phenol concentrations (EPC) [
186]. The results showed that gelam honey possessed lowest mic value against
S. aureus with a 5% (w/v) mic and mbc of 6.25% (w/v). The highest mic values were shown by pineapple honey against
E. coli and
P. aeruginosa as well as acacia honey against
E. coli with a 25% (w/v) mic and a 50% (w/v) mbc values. Agar inhibition assay showed kelulut honey to possess highest total antibacterial activity against
S. aureus with 26.49 EPC and non-peroxide activity of 25.74 EPC. Lowest antibacterial activity was observed in acacia honey against
E. coli with total activity of 7.85 EPC and non-peroxide activity of 7.59 EPC.
The antimicrobial efficacy of conventional antiseptics has been reported against
S. aureus (including MRSA),
Enterococcus faecalis (including vancomycinresistant
Enterococcus),
Streptococcus pneumoniae,
E. coli,
P. aeruginosa,
Clostridium perfringens,
Haemophilus influenzae and
Candida albicans using PVP-iodine, triclosan, chlorhexidine, octenidine and polyhexanide as conventional antiseptics used in wound treatment [
187]. The results showed that for chlorhexidine, octenidine and polyhexanide, mic
48 and mbc
24 ranged from 16 to 32 mg/L. Maximum values for triclosan ranged from 256 to 512 mg/L, with an efficacy gap against
P. aeruginosa, while the maximum values of PVP-iodine were 1024 mg/L, with a gap against
S. pneumoniae. Comparing the minimal effective concentrations, octenidine was the most effective. After 1 min, only octenidine and PVP-iodine fulfilled the requirements for antiseptics. We can conclude that, according to those reported data, honey shows mic and mbc values in the same order of magnitude than conventional antiseptics and potentially applicable after validation in clinical settings. As an example, commercially available dressings based on chemically treated honey are nowadays available in Europe [
188].