OUR EXPERIENCE WITH THE USE OF 40% BENZOIC ACID FOR NECRECTOMY IN DEEP BURNS

INTRODUCTION

Chemical necrectomy has been used for more than 33 years (in approximately 6000 patients) successfully at the Department of Burns and Reconstructive Surgery of the University Hospital in Brno. There is not much data available in Czech or worldwide literature about this method and there is even missing data from any serious analysis of clinical efficiency, comparison with other necrectomy methods and mainly possible complications for the patient.^1,2 Ointment with benzoic acid in white vaseline is applied on demarcated deep burns at the earliest on the sixth day after the burn. Layer of benzoic acid in white vaseline is applied on tulle de grass, which has a size approximately as the burn surface area and it is placed on the wound. It is fixed with several metal clips. The actual necrectomy is performed 48 hours later, usually under general anaesthesia (according to the extent). Necrosis either falls off with the dressing or after easy release with blunt instruments. The base of the wound is usually clean, there is mild bleeding and it is possible to apply a skin graft. The ointment is applied in one stage maximally on 10% of body surface area (see Figure 1–3). Toxicity of benzoic acid for human is generally very low. There has been only irritation of skin with pseudoallergic rash and eye damage reported.³ Literature reports that the minimal level of benzoic acid associated with toxic symptoms in human is 6.5 mmol/l.^4,5 Benzoic acid is completely absorbed from the gastrointestinal tract, it is incompletely absorbed through intact skin. Absorbed benzoic acid is relatively quickly metabolized in the liver by conjugation with glycine producing hippuric acid, which is then excreted by urine. There is no accumulation in the body.⁶ We have performed a clinical study with the goal to monitor the dynamics of absorption and elimination of benzoic acid applied on burned skin. This study documents how fast and in what extent there is absorption of benzoic acid through burned skin taking place, what plasmatic levels are achieved, how fast it is metabolized to hippuric acid, which is excreted in urine. We collected urine and calculated the quantity of excreted hippuric acid and correlated these calculations with plasmatic concentrations of both acids and extent of absorption surface and we also investigated, whether conjugation of benzoic acid with glycine producing hippuric acid does not lead to reduced plasmatic levels. Supplementary part of our study was monitoring of microbial colonization of areas after chemical necrectomy.

MATERIAL AND METHODS

There were totally 22 patients aged 6–85 years with skin burn IIb–III degree studied. There was 40% benzoic acid in white vaseline applied for 48 hours on burn area of 0.5–10% total body surface area. Venous blood was collected in the first 10 patients before application (0 minutes) and in 10, 20, 60 minutes and then 2, 4, 6, 12, 24 and 48 hours after application on burn area in order to determine benzoic acid, hippuric acid and glycine in serum. In the subsequent patients, when we knew the rough dynamics of absorption, we reduced the frequency of blood collections to half and we omitted collections in 10 and 20 minutes and after 2 and 4 hours. Urine for determination of hippuric acid was collected in four 12hourly intervals. Examination took place in the laboratories of the Department of Clinical Biochemistry, University Hospital in Brno-Bohunice. Determination of benzoic acid and hippuric acid in serum and hippuric acid in urine was performed using HPLC method – high performance liquid chromatography on HPLC Waters 2695–Alliance with diode array detector Waters 2996. Automatic analyser of amino acids Biochrom 20 (Biochrom-Pharmacia) was used for determination of glycine in serum.^7,8 Collections were performed according to the same time schedule as in case of benzoic and hippuric acid.

In all 22 patients were performed imprints or smears before chemical necrectomy for microbiological examination of the presence of potentially pathogenic microorganisms. The same was performed after removal of necroses 2 days later. (Fig. 1, 2, 3.)

RESULTS

1. Serum concentration of benzoic acid

The values in Chart 1 show that there is a quick rise of benzoic acid level in serum within the first hours after application. Most frequently measured maximal serum levels were 1–2 hours after application of ointment. Therefore we did not proceed with collections in the first hour in patients No. 11–22 and we also skipped the collections after four and six hours. In some patients we also collected sample after 72 hours; the levels were usually lower than the initial levels, therefore we also skipped them. Next stage of benzoic acid application on residual necroses followed in some patients after 48 hours and we either did not perform collections, or, if these were performed, concentration raised quickly again. Next 12 patients were already examined according to a reduced scheme.

We notice a relatively large difference between arithmetic means and median. It is due to the fact that the range of measured absolute concentrations is 0.008 mmol/l to 1.3 mmol/l, which is mainly dependent on the extent of surface with applied acid. Therefore, while median in 2^nd hour is 0.08 mmol/l, arithmetic means is severely biased by two patients with the highest levels (patient 6 and 9, the maximum of whom is around 1.0 mmol/l) and reaches the level of 0.226 within the same time. Curves and courses are relatively coherent in all patients.

2. Serum concentration of hippuric acid

Similarly to benzoic acid, also hippuric acid was examined in the first 10 patients more frequently and in the remaining 12 according to a reduced scheme (Chart 2). Measured variability of absolute serum concentrations must have some cause. The most logic hypothesis is that patients with higher extent of applied ointment will have higher concentrations. Table 1 summarizes basic facts about each patient.

3. Plasma concentrations of glycine

Serum concentrations of glycine during application of benzoic acid are quickly declining in the first hours after application, which demonstrates consumption of glycine for conjugation with benzoic acid to produce hippuric acid (Chart 3). All patients have initially higher levels than after 48 hours, but none of them demonstrated decline below the lower reference range. Reference range for glycine in adults is 120–554 µmol/l and in children 117–223 µmol/l.

4. Total absorbed quantity of benzoic acid during chemical necrectomy

The total absorbed and excreted quantity of benzoic acid may be theoretically calculated from collected urine, while we assume that 1 molecule of hippuric acid developed from 1 molecule of benzoic acid. The total amount of excreted hippuric acid is then multiplied by molecular weight of benzoic acid (M = 122) and we obtain an approximate quantity of absorbed acid. It should be noted, however, that part of benzoic acid is metabolized by glucuronization, i.e. another, minor pathway. Furthermore, not all benzoic acid in blood and urine is due to chemical necrectomy, since it is widely used as a preservative in cosmetics, food and drinks. One mol of hippuric acid produces 1 mol of benzoic acid and if multiplied by molar concentration of benzoic acid, which is 122 g/mol, we obtain total weight of benzoic acid, from which hippuric acid developed: m = n x M

Results after 48 hours are listed in the last column of Table 1. However, in comparison with serum concentrations, the quantity of applied benzoic acid correlate only little and seem to be very inaccurate and unsuitable for possible monitoring of safety of chemical necrectomy. Collection of urine is demanding with regards to good cooperation of the patient and medical staff and there is more space for failures and inaccuracies. This examination could not be performed at all in 5 patients.

5. Microbiological examination of areas after necrectomy

Imprints for culture before application of benzoic acid and after 2 days were collected in all patients. No microbes were cultured in most patients or their quantity declined. Only in 3 patients the finding worsened and there was positive culture of coagulase negative Staphylococcus or once higher count of Staphylococcus aureus.

DISCUSSION

Absorption and elimination of benzoic acid

Obtained results indicate that benzoic acid is absorbed during chemical necrectomy as performed by our method. It is understandable, since the main barrier of skin is stratum corneum, which is completely missing in burn skin. The quantity of absorbed acid depends mainly on the extent of the area, on which it is applied as shown in Chart 4. This shows maximal levels of benzoic acid in patients up to 4% of applied benzoic acid. Concentrations range from 0.008–0.084 mmol/l and very well correlate with the extent of absorption area. In case of greater extents of absorption area, the variability of serum concentrations is already higher but ranges in the extent from 0.125–1.3 (Chart 5). Peak plasmatic concentration of benzoic acid may be moved to later hours in patients with thicker layer of necrosis. Our explanation is that there is also thermal damage to the subdermal vascular plexus and since subcutaneous tissue is not well vascularized, there is slower and apparently also incomplete absorption of benzoic acid taking place. Metabolism of benzoic acid to hippuric acid depends on good liver function. Excretion of hippuric acid is then influenced by kidney function.

In our group we demonstrated 2 types of absorption and metabolism of benzoic acid and divided them into two groups. The first group includes patients, who quickly metabolize benzoic acid to hippuric acid and levels of hippuric acid exceed concentration of benzoic acid in each sample. This type of metabolism correlates mostly with uncomplicated course and successful chemical necrectomy. This group included 15 patients (patient No. 2, 3, 5, 8, 10, 11, 12, 13, 14, 15, 16, 17, 19, 20, 21). The second group included patients in whom the ratio of both acids is the opposite (patient No. 1, 4, 6, 7, 9, 22). Common feature is higher age, greater total extent of the burn, development of burn shock or possible co-morbidities. The cause of this feature is not very clear. It should not be a lack of glycine with regards to the aforementioned results. With increasing plasmatic concentration of benzoic acid rises also concentration of hippuric acid. However, at some point, it seems as if there was conjugation capacity of hepatocytes exceeded and even in spite of high level of benzoic acid, concentration of hippuric acid does not continue to rise and starts to decline. In spite of that, however, there continues rapid decline of plasmatic concentration of benzoic acid. Is there a different route of elimination used or benzoic acid is excreted to urine in unchanged form? Reduced capacity of the liver to produce adequate quantity of hippuric acid could be due to a lack of ATP, which is apparently a common feature in patients, who suffered burn shock. This however does not explain continuing rapid elimination of benzoic acid from plasma.

Mechanism of action on necrosis, histology

The exact mechanism of action of benzoic acid during chemical necrectomy has not been clarified yet. There were initial thoughts that there is some kind of necrolysis, i.e. caustic effect on necrotic part of burn skin. It seems, however, from clinical observation and several collected histological samples that benzoic acid acts as a strong chemoattractant to immunocompetent cells, which intensively and rapidly migrate to the layer between vital and non-vital tissues and by the production of colagenases and proteases cause release of the bonds between necrosis and undamaged tissue. The principle is therefore based on significant acceleration of physiological inflammatory reaction, when the body after several weeks is able to remove non-vital tissue alone. When lifting the necrosis after application of benzoic acid, we observe collection of “pus–like” fluid, which is however mostly sterile. It is logical with regards to the anti-microbial and anti-mycotic effect of benzoic acid and verified by routine practice of collecting microbial imprints from areas after necrectomy. The following cellular components contribute on the release: neutrophils – polymorphonuclears, macrophages producing collagenases, metalloproteinases, interleukins and cytokines. Possible predictive factors of failure of this method seem to be generally poor medical condition leading to induced immunosuppression of the organism, which clearly does not respond to an immune stimulus, and also thermal destruction of skin reaching to subcutaneous fat. Our hypothesis is based on the fact that in case there is damage to subdermal vascular plexus, then immunocompetent cells penetrate slower to less perfused fatty tissue and the method of chemical necrectomy for this type of damage is not effective.

We attempted to compare the efficiency of chemical and classical tangential necrectomy with the following conclusions (Table 2).

CONCLUSION

Efficiency of benzoic acid during necrectomy of burns according to our long-term clinical practice and performed study is characterized as follows:

• 40% benzoic acid in white vaseline is effective in most skin necroses caused by all types of burns. Healing of grafts on wound bases after chemical necretomy is the same – almost hundred percent – as after all types of sharp necrectomy
• The main advantage of necrectomy with benzoic acid is its selectivity and significantly lower blood losses.
• The disadvantage is moderate pain several hours after application of benzoic acid and efficiency at the earliest 6 days after burn. It is less successful or unsuccessful in patients who are immunocompromised due to any reason.
• Our study demonstrated absorption of benzoic acid through skin necrosis, however its level in blood remains always low and below the toxicity level – i.e. it is a safe method.
• There was antibacterial and anti-mycotic effect of benzoic acid on wound area demonstrated. In most patients there were no microbes or their quantity declined in the area after chemical necrectomy.
• The method is suitable mainly at suboptimal clinical conditions, where it is not possible to provide perfect sterility and sufficient amount of blood derivatives to supplement blood losses.
• Very suitable indication for chemical necrectomy is the use in the elderly and polymorbid patients, since it is possible to avoid general anaesthesia in patients during removal of necroses.
• This method seems advantageous in some areas, such as dorsum of the hands, feet, pretibial area, where tangential necrectomy could result in damage of deep structures.
• In case of extensive burns, we consider to be optimal combination of benzoic acid with other types of necrectomy, which enables removal of necrosis from the patient in shorter period of time with less blood loss.

Corresponding author:

Zuzana Jelínková, M.D.

Department of Burns and Reconstructive Surgery

Medical Faculty of Masaryk University and University Hospital Brno

Jihlavská 20

625 00 Brno

Czech Republic

E-mail: jelinkova.zuzana@fnbrno.cz