Proteolytic Enzymes and Wound Debridement: A Literature Review

Surgery, burns, and injuries often result in wounds that exhibit poor or incomplete healing. Chronic wounds impact the quality of life of nearly 2.5% of the US population, with a higher prevalence among the elderly. These wounds often lead to complications and contribute to significant health care costs.¹ Unhealed wounds also significantly affect the patient’s daily life, affecting their physical, mental, and social well-being.²

A wound is defined as any damage to the physiological or anatomical structure of the skin, leading to the loss of functionality in the affected area.³ Wounds are categorized as acute, which are capable of healing,⁴ and hard-to-heal, in which the healing process does not progress through the typical stages of healing in a timely manner.⁵ Due to delayed or discontinued treatment, as well as intrinsic and external factors, a wound may persist in a constant state of inflammation. This could potentially lead to issues such as pain, infection, scarring, disability, amputation, or even death.^2,6,7

Various factors, such as systemic diseases, diabetes, and aging, significantly contribute to the complexities of wound healing.^2,8 The creation of necrotic tissue poses a major challenge in the wound healing process, hindering healing and often leading to microbial contamination.⁹ In clinical practice, surgical debridement is commonly performed to “clean” wounds of necrotic tissues. However, this essential procedure is invasive and can impede the healing process. A noninvasive and effective method of nonsurgical debridement could make a substantial contribution to the enhancement of wound healing.

The electronic databases PubMed (National Library of Medicine) and Google Scholar were searched for English-language articles published from 1974 to 2023. The keywords searched were “enzymatic debridement”, “wound healing”, “collagenase”, “bromelain”, “proteolytic enzymes”, and “debridement”. 

In wound management, debridement is the process that eliminates attached, dead, or infected tissue from the wound. It is distinct from cleansing, which involves the removal of impurities, metabolic wastes, or foreign materials.¹⁰ There are various debridement methods, and the selection of the most suitable depends on tissue type, pain, age of the wound, practitioner skills, resources, regulations, and the patient’s needs. The debridement types discussed in the current article are autolytic, larval, mechanical, sharp (or episodic), surgical, and enzymatic. 

Autolytic debridement

Autolytic debridement enhances the wound environment through the use of bandages suitable for wound dressing.^10,11 These bandages either add moisture to or remove it from the wound area, facilitating the body’s autolytic process whereby enzymes dissolve dead tissue. Autolytic debridement is painless, noninvasive, and easy to perform, but it is slow and may potentially increase the risk of infection.^10,11

Larval debridement

Current larval debridement practice involves the use of digestive secretions and excretions from larvae of Lucilia sericata. These larvae use proteolytic enzymes to liquefy and digest dead tissue, kill bacteria, and stimulate wound healing. Although this treatment provides rapid and selective removal of dead tissue, it can be costly and may not be easily accepted by all patients.^12-14

Mechanical debridement

Mechanical debridement swiftly removes dead wound tissue, but it can be painful.¹² It involves placing wet gauze over the wound, then allowing it to dry and adhere to the wound surface before pulling the gauze away.¹⁵ The monofilament wound debridement pad Debrisoft (Lohmann & Rauscher USA, Inc.), a less painful mechanical removal method, has been in use since 2011.¹⁶ This gauze is used to gently wipe the wound to remove exudates, dead cells, and wound debris.¹⁶ 

Sharp (or episodic) debridement

Sharp (or episodic) debridement has been shown to be highly beneficial for wound closure. It helps by removing necrotic tissue, bacteria, and other foreign materials from the wound, which can significantly accelerate the healing process and enhance the effectiveness of other therapeutic measures.^17,18 Dead tissue is removed using scissors, a scalpel, and/or forceps in sharp debridement. It is crucial that the practitioner differentiate between viable and nonviable tissue. This debridement method can be painful for the patient, and if the practitioner is not adequately trained, there is a risk of either leaving behind dead tissue or removing healthy tissue. Another limitation lies in the practice of sharp debridement outside a hospital setting, which may carry an increased risk of microbial contamination.^12,15

Some advanced sharp debridement techniques utilize mechanical devices to remove dead tissue. These methods can offer greater precision and efficiency compared with manual sharp debridement.^19-21 Various types of powered debridement exist, including tangential hydrosurgery^19,20 and ultrasound-assisted debridement.²¹

Tangential hydrosurgery uses a high-pressure stream of sterile saline to selectively remove nonviable tissue while preserving healthy tissue.¹⁹ It is a rapid method that can be painful, depending on the patient. The expense of the device contributes to the overall cost of hydrosurgery. Tangential hydrosurgery is reported to minimize mechanical trauma and to reduce the risk of damaging healthy tissue.^19,20

In contrast, ultrasound-assisted debridement uses ultrasonic energy to break down necrotic tissue. The ultrasonic waves create cavitation bubbles that disrupt and liquefy necrotic tissue, making it easier to remove. Ultrasound-assisted debridement is minimally invasive and can be more comfortable for the patient, reducing the likelihood of pain and minimizing damage to surrounding healthy tissue.²¹

Surgical debridement

Surgical debridement is the most common method and requires an operating room and a surgeon. This method provides immediate results with the complete removal of damaged tissue, but it is a traumatic experience often requiring anesthesia, and it is associated with bleeding, heat loss, and nonselective removal of healthy tissue. Wound expansion is linked to microbial infection, and some patients may not tolerate the process due to underlying diseases.^{9,12,15,18,22}

Enzymatic debridement

Enzymatic debridement involves applying to the wound a preparation containing a proteolytic enzyme with precise specific activity per gram. This method is used to remove necrotic tissue from chronic skin wounds and burns.^23,24

Enzymes are proteins that serve as the primary catalysts in biological systems. Proteolytic enzymes are those that hydrolyze peptide bonds, contributing to the degradation of damaged cells and necrotic materials. Moreover, they play a role in restricting edema, absorbing hematoma, and reducing wound pain.²⁵

Aspartic or aspartyl peptidases, which are characterized by 2 chains of aspartic acid, are acidic endopeptidases. They exhibit activity within a pH range of 3 to 6 and with an isoelectric point of 3 to 4.5, and they are inhibited by pepstatin A in the presence of copper(II) ions. These enzymes are produced by animals, plants, bacteria, fungi, and viruses.²⁶

Cysteine or thiol peptidases function as both endopeptidases and exopeptidases, relying on a cysteine residue.²⁶ Their activity is observed at pH 4.5 to 7.²⁶ The histidine and cysteine duo is present in all thiol peptidases, with well-known examples being papain, ficin, and bromelain.²⁷

Serine peptidases, featuring 1 serine at the active site, act as both endopeptidases and exopeptidases. They reach maximum activity at pH 6 to 11, with an isoelectric point of 4 to 6. The catalytic triad involves histidine, serine, and aspartate. Prominent examples of serine peptidases include chymotrypsin, trypsin, elastase, thrombin, plasmin, and subtilisin.^26,27

Metallopeptidases, which function as both endopeptidases and exopeptidases, depend on ions (zinc, cobalt, manganese, nickel, copper, and iron) for catalysis. The active site is represented by histidine–glutamic acid–any amino acid–any amino acid–histidine. Carboxypeptidases, aminopeptidases, and dipeptidases (exopeptidases), along with collagenase (endopeptidase), are examples of metallopeptidases.^26,27

Threonine peptidases are characterized by the N-terminal threonine residue crucial for catalytic activity. The hydroxyl part of threonine serves as a nucleophile with catalytic function.²⁶

Glutamic peptidases, which form catalytic centers with serine, glutamic acid, and aspartic acid, fall under the classification of serine peptidases. They exhibit activity at pH 2 and pH 1.1, using casein and hemoglobin as substrates, respectively.²⁶

Asparagine peptide lyases, unlike hydrolytic proteolytic enzymes, act solely through self-cleavage occurring on the C-terminal side of the asparagine residue’s active site.²⁶

Enzymes used for wound debridement

The various enzymatic agents used in attempted clinical management of wound debridement are listed in the Table. 

The first commercially available enzymatic debridement substance was sutilains ointment (Travase), derived from Bacillus subtilis. Sutilains ointment was introduced in the 1960s²⁷ and was clinically tested on a large number of patients.²⁸ While some authors drew a positive conclusion about its efficacy as a debriding agent,²⁹ others did not share this assessment.³⁰ In subsequent decades, it became widely recognized as a useful enzymatic debridement agent. However, an increase in wound infections was observed, potentially due to the moist environment required for its application, which seemed to foster bacterial growth.²⁹ To mitigate infection risk, simultaneous use with antimicrobial agents such as silver sulfadiazine was recommended at the time.²⁹ Moreover, adjustments in application frequency and timing postburn could hasten the debridement process, allowing for quicker wound closure through autologous skin grafting.³⁰ Despite its effectiveness and popularity in American burn units, this product was discontinued in the 1990s.²⁹

Currently, proteolytic enzymes are used for burns and chronic ulcers. Collagenase and bromelain are commonly used proteolytic enzymes for wound debridement today, while papain, which was more widely implemented during the 1980s and 1990s, is still in use in certain countries around the world.

Collagenase

Collagenase is a proteolytic enzyme that is typically derived from Clostridium histolyticum^24,31 or Vibrio alginolyticus.³² It possesses the ability to hydrolyze collagen and other proteins, which are key components of necrotic tissue.^24,33 By breaking down collagen strands, collagenase dislodges debris and aids in the removal of necrotic tissue from the wound surface.^24,33 Collagenase is available under different brand names, such as Collagenase Santyl Ointment (Smith & Nephew), Hyalo4 Start (Fidia Farmaceutici S.p.A.), Bionect Start (Fidia Farmaceutici S.p.A.), and Iruxol N or Mono (Smith & Nephew), and it is used for debriding necrotic tissue in chronic cutaneous wounds and burns.^24,32,34 Collagenase is the most widely used debriding enzyme globally, and it is estimated that in 2021, 54,414 patients in the United States were treated with this enzyme, resulting in an estimated total of 164,891 prescriptions.³⁵

Collagenase lacks antimicrobial properties, so its optimal use involves combination with a topical antibiotic to prevent infection.^24,31 Accelerated wound healing has been observed after the debridement of devitalized tissues, potentially leading to shorter hospital stays, reduced time for wound care, and decreased patient discomfort.³⁴ However, the enzyme’s activity is sensitive to the wound environment, temperature, and heavy metals—all factors that can reduce its effectiveness.^33,34 An associated side effect of collagenase is irritation of healthy skin, with some patients experiencing a stinging sensation, particularly if they are hypersensitive to the enzyme.²⁴ Various antimicrobial wound dressings, particularly those based on dye and collagen, have shown good compatibility with collagenase.³⁶

Many studies have touted clostridial collagenase ointment as a cost-effective method for treating chronic dermal ulcers.^34,37,38 However, a 2017 systematic review and meta-analysis conducted by Patry and Blanchette²³ concluded that most studies had a high risk of bias and varied outcomes. Patry and Blanchette²³ identified 4 RCTs comparing collagenase with placebo. The first RCT, published in 1969³⁹, compared collagenase 0.05% and a combination of collagenase 0.05% with neomycin against a placebo in 47 subjects with a total of 62 decubitus or vascular ulcers. Collagenase treatments resulted in complete debridement in 58 of 62 wounds, while placebo treatment achieved complete debridement in only 1 of 15 wounds. The authors of that RCT concluded that collagenase effectively debrided chronic dermal and decubitus ulcers.³⁹ A small study from 1973 involving 20 subjects examined collagenase with polymyxin powder vs. placebo for dermal and decubitus ulcers, showing significant decreases in wound size as well as in “pus, odor, necrosis, and inflammation” in the collagenase group (P < .01 and P < .07, respectively).⁴⁰ In 1975, a small RCT with 11 subjects and 28 wounds (primarily pressure ulcers) found improvement in 14 of 17 ulcers treated with collagenase, with no improvement in the placebo group.⁴² A small double-masked RCT⁴¹ (patients, n = 30) compared collagenase (Iruxol Mono) with a placebo in outpatients with various ulcer types, finding significant improvements in wound size reduction, debridement, and epithelialization in the collagenase group (P < .01 for each outcome), despite differences in wound characteristics and systemic antibiotic use in the placebo group.^39-42 

Moreover, in a retrospective observational study of hyaluronic acid and V. alginolyticus collagenase ointment involving 70 patients with chronic wounds of various etiologies, De Francesco et al³² found that the frequency of debridement efficacy in terms of wound bed cleansing increased from 26% after 2 weeks to 93% after 4 weeks. Complete healing was observed in 62 patients within an 8-week period. Overall operator and patient satisfaction after 8 weeks were 100% and 90%, respectively, with all patients reporting reduced pain. The investigators concluded that the combined action of hyaluronic acid and collagenase ointment reduced healing time while improving the quality of healing and decreasing pain.³²

Bromelain

Bromelain, which is recognized as a proteolytic and hydrolytic enzyme, exhibits a selective capacity for debriding heat-denatured proteins. Its commercial form, NexoBrid (MediWound), and its diluted form, EscharEx (MediWound), are medicinal products that incorporate, apart from bromelain, other proteases including escharase. These substances are commonly derived from the stem of the pineapple plant.^43-47

Some in vivo and clinical studies of bromelain mixtures have demonstrated rapid debridement or dissolution of necrotic skin.^48-50 Conversely, control gels (acting as hydration agents) showed no debridement, and the integrity of blood vessels, hair follicles, and porcine glands remained unaffected.^49,51 Two different studies of bromelain, a porcine study⁵¹ and an in vitro study of human cells,⁵² reported no adverse effects on normal or injured skin, underscoring its selectivity. In vitro studies have indicated that bromelain, particularly at elevated concentrations, may have a toxic effect on keratinocytes and fibroblasts.⁵² 

Products containing bromelain were first approved by the European Medicines Agency in 2012 and are commercially available globally.⁴⁶ Clinical and animal studies exploring the application of these products in various cases, including upper extremity burns,²² hard-to-heal wounds,^32,48,54,55 e-cigarette burns,⁴⁷ full-thickness facial burns,⁴⁸ cutaneous systemic sclerosis in mixed depth flame burns,⁵⁵ and genital burn injuries,⁵⁷ have demonstrated its safety, selectivity, and efficacy.^{48,50,53,54,56} 

Bromelain-based enzymatic debridement has also proven effective in preserving viable dermis, facilitating rapid healing with minimal scarring. Its advantages include immediate application upon hospital admission, reducing the risk of wound infections, and potentially eliminating the need for surgical escharotomy.^46,56,57 While pain medication or anesthesia may be necessary in some cases, the overall benefit of such debridement is cost-effectiveness, streamlining operating resources and time to healing.^46,58 However, it is worth noting that one study suggested that bromelain-based enzymatic debridement might be inadequate for managing burn wounds in patients with diabetes.⁵⁹ 

Several published studies have examined the efficacy of the concentrated form of bromelain-based debriding enzymes. In a single-center study, Cordts et al⁴⁶ found that bromelain-based treatment negated the need for further surgical intervention in 53.8% of patients with severe upper extremity burns. For those requiring surgery, the area needing skin grafts was reduced by 37%, although patients experienced prolonged wound closure of 28 days. Physical therapy commenced approximately 2 days postinjury.⁴⁶ Schulz et al⁴⁸ studied deep facial burns and demonstrated that bromelain enzymatic debridement significantly reduced the time to complete wound closure compared to traditional methods (19.85 vs. 42.23 days; P = .002). Additionally, fewer procedures were needed to achieve complete debridement (1.00 vs. 1.77; P = .003), and 77% of burns initially assessed as deep were found to be more superficial post-debridement. Moreover, the need for autografting was significantly reduced (15% vs. 77%; P = .002), and scar quality improved in several parameters.⁴⁸ In a multicenter RCT, Rosenberg et al⁵⁰ compared bromelain-based debridement with nonsurgical debridement and found the former to be associated with significantly decreased time from injury to complete debridement (2.2 vs. 8.7 days; P < .0001), reduced need for surgery (24.5% vs. 70.0%; P < .0001), reduced area of burns excised (13.1% vs. 56.7%; P < .0001), and reduced need for autografting (17.9% vs. 34.1%; P = .01). There were no significant differences between treatment groups in adverse events.⁵⁰ 

Interesting results have also been achieved with the diluted form of commercially available bromelain-based enzyme mixtures. In a multicenter RCT by Shoham et al,⁴⁴ the bromelain-based debridement group had a significantly higher rate of complete debridement compared to the placebo group (55% vs. 29%, P = .047).⁴⁴ Additionally, in a small clinical trial conducted by Snyder et al,⁶⁰ 7 of the 10 patients who completed treatment achieved complete debridement after a median of 2 applications of the bromelain-based treatment. By the end of the 2-week follow-up period, an average reduction in wound area of 35% was achieved. Furthermore, in patients who initially tested positive for biofilm, the biofilm was reduced to single individual or no detectable microorganisms by the end of treatment.

European consensus meetings have established preliminary guidelines for bromelain-based enzymatic debridement,^61,62 and several European hospital centers adhere to these recommendations in treating burn patients with such products.⁶² In certain Italian wound care units, enzymatic debridement has been particularly prioritized to reduce the need for surgical intervention and minimize blood loss, especially as part of the approach to managing care during challenging circumstances like the COVID-19 pandemic.⁶³

Papain

Papain, a proteolytic enzyme derived from the fruit of Carica papaya, is commonly used in enzymatic debridement due to its ability to digest nonviable tissue. Papain cleaves all proteins containing cysteine.^64,65 However, papain alone is relatively ineffective; activators are required to enhance its potency.⁶⁶ One such activator is urea, which acts by interfering with disulfide bonds in proteins, exposing sulfhydryl groups that activate papain.⁶⁶ The addition of copper chlorophyllin has reported in some cases.⁶⁴ The combination of papain and urea in debriding ointments has been shown to significantly increase digestive activity compared to papain alone.⁶⁴ This combination effectively denatures nonviable tissue, making it more susceptible to enzymatic digestion.⁶⁶ Additionally, papain-urea agents have been found to efficiently degrade necrotic tissue, excluding collagen and fibrous tissue.⁶⁶ However, concerns have been raised regarding its use, particularly its potential to continue digesting healthy cutaneous tissue after degrading necrotic tissue.⁶⁵ This can lead to significant inflammation, erythema, and discomfort during therapy, making papain-urea products potentially toxic for wounds,⁶⁵ especially in patients with major burns.⁶⁶

Some studies have used alternative enzymes such as chymotrypsin (derived from maggots), aurase, and Antarctic krill enzymes (Euphausia superba) (hereafter krill), with most studies yielding inconclusive results.

Chymotrypsin derived from the larvae of Lucilia sericata is resistant to endogenous wound protease inhibitors but seems to be restricted by blood plasma inhibitors.⁶⁷ The larval secretion consists of trypsin-like and chymotrypsin-like proteases and metalloproteases that stimulate fibroblasts and aid in bacterial disinfection.⁶⁷ 

Maggot-derived chymotrypsin I, which is biochemically distinct from human α-chymotrypsin, demonstrates resilience in the wound environment due to its lack of inhibition by wound eschar.⁶⁸ This characteristic enables it to contribute effectively to wound debridement during larval biotherapy.⁶⁸ Further supported by in vitro studies, the use of insect-derived products such as maggot-derived chymotrypsin I is suggested to be beneficial not only for debridement but also for the stimulation of granulation tissue formation.⁶⁷ However, the application of such enzyme-based treatments requires precise control over dosage to maximize therapeutic benefits while minimizing potential adverse effects. Larval chymotrypsin appears promising for further study due to its debridement properties.^67,68

Aurase is a novel recombinant proteolytic enzyme derived from maggot saliva.⁶⁹ It demonstrates selectivity for fibrin, and at higher concentrations it exhibits additional proteolytic action against collagen and elastin. In a methicillin-resistant Staphylococcus aureus biofilm model, the enzyme, formulated in a proprietary gel, significantly reduced bacterial counts and improved wound vascularity, supporting its potential as an effective debriding agent.⁶⁹ 

Toxicology studies on aurase have shown no observable systemic effects, even when administered in high concentrations via bolus intravenous injection.⁶⁹ This absence of systemic adverse events is reported to be associated with aurase’s rapid binding to α-1 antitrypsin, an endogenous protease inhibitor, forming a high-affinity complex, potentially covalent. This interaction suggests that aurase selectively targets necrotic tissue for digestion,⁶⁹ a property that may lead to deactivation upon contact with blood. Such characteristics position aurase as a promising agent for the specific treatment of necrotic tissue, minimizing the risk of damage to healthy cells.

Krill possesses endopeptidases and exopeptidases, ensuring nearly complete breakdown of proteinaceous substrates to amino acids. Applied twice a day at a concentration of 3 U/mL, krill enzyme treatment appears to be an effective debridement method in animal wound models.⁷⁰ In vitro studies have highlighted the superior wound debriding efficacy of krill enzymes when compared to fibrinolysin/DNAse, trypsin, collagenase, and streptokinase and streptodornase.⁷¹ Nonetheless, applying these findings to real-life scenarios could be complicated by serine protease inhibitors present in serum and wound fluid, potentially hindering the activity of the enzymes.⁷¹ 

A clinical trial involving 31 patients with venous leg ulcers assessed the debriding potential of a preparation derived from krill against a routine nonenzymatic treatment.⁷⁰ Five milliliters of a solution of 6 U/mL of krill enzymes applied twice a day for 7 days to venous leg ulcers showed the removal of necrotic tissue. The krill enzyme group experienced a 53% reduction in necrotic area within 7 days, compared to no reduction in the control group. The median time for ulcer cleaning was shorter in the krill enzyme group (7 days) than in the control group (10 days), and subsequent autologous skin grafts yielded similar success rates in both groups, with no observed side effects. This led to the conclusion that krill enzymes are an effective option for the debridement of venous leg ulcers.⁷⁰

Kiwifruit (Actinidia deliciosa), which is rich in the protein-dissolving enzyme actinidin (thiol protease), proved highly efficient in an in vivo enzymatic debridement study in rats, offering an alternative debridement enzyme.⁷² Interestingly, debridement and scar contraction occurred more rapidly in the group treated with kiwi actinidin, occurring in nearly half the time compared with the untreated group. Following swift enzymatic debridement, healing progressed as expected, with no signs of harm or toxicity to the surrounding healthy tissue.⁷² 

Dispase, a neutral metalloprotease produced by Bacillus polymyxa, has shown significant promise in the field of wound healing and debridement, particularly in the aftermath of cryosurgery-induced ulcers. Its efficacy, highlighted in a pilot comparative clinical trial, underscores the enzyme’s potential for broader clinical application.⁷³

Characterized by its selective proteolytic activity, dispase effectively cleaves type IV collagen and fibronectin, key components in epithelial–mesenchymal interactions; thus, use of dispase may facilitate the selective removal of damaged tissue without harming the surrounding viable tissues. This selectivity, coupled with the observed absence of side effects in both animal models and clinical trials, positions dispase as a safe and effective wound healing agent. In a recent study, its experimental application not only prevented the formation of necrotic tissue—a stark contrast to outcomes observed with octenidine gels—but also promoted a faster healing process, attributed in part to the zinc content of dispase that may enhance fibroblast mitosis.⁷³ Given these promising results, further investigation into dispase’s utility in routine clinical settings is both justified and necessary.

Ozcan et al⁷⁴ compared the effectiveness of debridement using C. histolyticum–derived collagenase ointment vs. surgical excision, comparing 78 children treated with collagenase and 41 subjects treated with surgical excision. Treatment allocation was not randomized, and decisions were made based on clinical judgment. The study authors indicated that the time required for complete removal of necrotic tissue did not differ significantly among the 3 treatment methods (collagenase alone, collagenase plus surgical management, or surgical management only). Patients treated with collagenase alone were less likely to require a blood transfusion compared to those treated with collagenase plus surgical intervention or those managed solely by surgery. Patients originally treated with collagenase were also less likely to require surgical excision compared to those managed exclusively by surgery. Moreover, patients treated with collagenase had a shorter hospital stay compared to those managed solely by surgery or those treated with collagenase followed by surgical intervention.⁷⁴

In a small clinical study, it was observed that a formulation containing a combination of hyaluronic acid and collagenase from V. alginolyticus demonstrated significantly greater debridement than surgical treatment after 4 weeks.⁷⁵ However, that study did not identify similar efficacy of C. histolyticum–derived collagenase.

In 2017, Schulz et al²² discovered that enzymatic debridement exhibited superiority over surgical debridement in the treatment of severely burned hands. Bromelain was used for the enzymatic debridement process. This approach significantly reduced healing time, facilitated the earlier removal of necrotic tissue, and did not affect burn depth or skin structure. Three months after debridement, there was no significant difference in scar quality between the treatment groups.²²

In another study by Schulz et al,⁴⁸ enzymatic debridement outperformed surgical debridement for deeply burned faces. Bromelain was used for eschar removal in this study as well. The enzymatic method was deemed safe and more selective. Compared to the surgical approach, a shorter period was required for the completion of healing and treatment processes. Nonetheless, 77% of wounds in the enzymatic group healed without the need for skin grafts. After 12 months, the enzymatic group exhibited superior scar quality and aesthetic outcomes, particularly in terms of pigmentation, thickness, relief, pliability, surface area, stiffness, and scar irregularity.⁴⁸

Most studies have compared enzymatic debridement with the standard of care or the absence of debridement. However, some studies have compared different proteases. 

Clinical trials and in vitro studies comparing V. alginolyticus and C. histolyticum collagenases revealed differences in their aggressiveness. V. alginolyticus exhibits physicochemical properties that make it user-friendly, less painful, and nontoxic to healthy tissue.^75,76 In brief, Onesti et al⁷⁵ observed a significantly better debriding capacity of a formulation containing a combination of hyaluronic acid and collagenase from V. alginolyticus compared to the standard formulation with collagenase from C. histolyticum at 4 weeks. Additionally, in an in vitro study Di Pasquale et al⁷⁶ demonstrated that the formulation containing hyaluronic acid and collagenase from V. alginolyticus was significantly more selective for collagen fibers in the wound area, and therefore on the necrotic tissue, compared to the standard formulation with collagenase from C. histolyticum. Consequently, the formulation containing hyaluronic acid and V. alginolyticus–derived collagenase appeared not to affect the extracellular matrix of the wound area.⁷⁶

Research on enzymatic debridement agents has compared the efficacy of C. histolyticum–derived collagenase and papain-urea formulations in wound management. Collagenase has demonstrated the capability to degrade essential wound components such as collagen and elastin both in vitro and clinically, proving its effectiveness in the breakdown of wound matrices.⁷⁷ In contrast, papain-urea formulations have shown a stronger performance in degrading fibrin and collagen and have been particularly noted for their efficiency in pressure injury debridement.^78,79 Additionally, studies have suggested that papain-urea not only excels in reducing nonviable tissue but also enhances granulation tissue formation more effectively than collagenase does. For instance, an RCT of lower extremity chronic wounds in 100 patients highlighted that treatment with papain-urea in a hydrophilic ointment led to more visible necrotic tissue reduction and improved granulation during weeks 2 to 4 of treatment, as compared to a collagenase formulation in a petrolatum base, although no significant differences were noted in overall healing rates.⁷⁸ 

The choice of vehicle in enzymatic ointments has also been underscored as a critical factor influencing the debridement efficacy of the ointment. Papain-urea, which typically is found in a hydrophilic ointment base, has been shown to facilitate better enzyme release and absorption into the wound site compared to the more hydrophobic petrolatum base used for C. histolyticum collagenase.⁷⁹ This property may contribute substantially to the observed differences in clinical outcomes. Furthermore, comparative studies have consistently reported that papain-urea not only more effectively reduces slough and necrotic tissue but also promotes faster and more robust granulation tissue development compared with collagenase. This superior performance in both enzymatic debridement and facilitation of wound healing processes underscores the potential of papain-urea as a preferable choice in clinical settings in which rapid and effective wound debridement is crucial.²⁴

Significant findings also emerged from a research study comparing the effectiveness of bromelain compared with C. histolyticum collagenase for eschar removal from experimental porcine burns.⁸⁰ The study authors observed that all full-thickness burns treated with bromelain-enriched products experienced complete eschar removal after a single application, in stark contrast to those treated with collagenase, with none of the latter demonstrating complete eschar removal even after 14 days of treatment.⁸⁰ Similarly, deep partial-thickness burns treated with bromelain exhibited complete eschar removal after only 1 application. In comparison, deep partial-thickness burns treated with collagenase showed no complete eschar removal within the first 10 days. These results led to the conclusion that bromelain-enriched debriding agents may facilitate quicker eschar removal than debriding agents containing collagenase.⁷⁷

While this review extensively covers various aspects of debriding enzymes, there are inherent limitations associated with the type of literature and the dynamic nature of the field. First, as a narrative review and not a meta-analysis, this review does not statistically analyze the data from the studies included, which may limit the ability to draw definitive conclusions about the effectiveness and safety profiles of different enzymes. Second, the literature on debriding enzymes, although vast, often contains studies that provide limited information. Many articles do not fully elaborate on experimental conditions, dosages, and long-term outcomes, while others analyze the effect of enzyme mixtures with other substances without reporting effectiveness of the enzyme alone, which can obscure comprehensive comparisons and conclusions. Finally, research on debriding enzymes is an actively evolving area in which new findings continually reshape health care professionals’ understanding. As global knowledge expands and new research methodologies are developed, previously established findings may be challenged or refined. Thus, conclusions drawn today may need to be reevaluated in the future considering new evidence. 

The evaluation of debriding methods and means necessitates a comprehensive analysis encompassing safety, selectivity, efficacy, speed, and cost efficiency. The clinical value of a debridement approach is significantly enhanced when it minimizes systemic side effects, avoids damage to surrounding healthy tissues, ensures complete and swift debridement, and is both cost-effective and simple to use.

Enzymatic wound debridement is recognized for its efficacy, sparing healthy tissues while accelerating the healing of traumas. Debridement using bromelain has been shown to facilitate the rapid removal of necrotic tissue with minimal adverse effects. Particularly, collagenase sourced from V. alginolyticus has been effective in debriding chronic wounds, thereby hastening the healing process. The efficacy of debridement can be significantly enhanced through the combination of enzymes with other substances, such as papain with urea, and collagenase from V. alginolyticus with hyaluronic acid. Furthermore, research involving other enzymes, including those derived from E. superba, chymotrypsin I, aurase, actinidin, and dispase, has demonstrated improved debridement outcomes.

Overall, proteolytic enzymes have emerged as a promising alternative to surgical debridement, offering a potentially safer and more effective solution. However, despite their advantages, enzymatic debridement methods remain underutilized and underexplored. Further research is needed to investigate various enzymes and their effectiveness compared with traditional surgical techniques. Such studies not only have the potential to broaden the scope of viable debridement options but may also help improve patient outcomes by providing safer, quicker, and more cost-efficient debridement methods.

The authors are grateful to Mrs Anastasia Fronimaki for her assistance in the correct use of the English language in the manuscript.

Authors: Evdoxia Mathioudaki, PharmD, MSc^1,2; Andreas Vitsos, PharmD, MSc, PhD²; and Michail Christou Rallis, PharmD, MSc, PhD²

Affiliations: ¹Farmeco Dermocosmetics SA, Research & Development Department, Athens, Greece; ²Department of Pharmacy, National and Kapodistrian University of Athens School of Health Sciences: Ethniko kai Kapodistriako Panepistemio Athenon, Zografou, Greece

ORCID: Rallis, 0000-0002-3802-197X; Vitsos, 0000-0002-2246-5381

Disclosure: The authors disclose no financial or other conflicts of interest.

Correspondence: Michail Christou Rallis, PharmD, MSc, PhD; National and Kapodistrian University of Athens, School of Health Sciences, Pharmaceutical Department, Section of Pharmaceutical Technology, Panepistimiopolis, Zografou 157 84 Greece; rallis@pharm.uoa.gr

Manuscript Accepted: July 31, 2024