|Year : 2014 | Volume
| Issue : 2 | Page : 38-46
Does wound pH modulation with 3% citric acid solution dressing help in wound healing: A pilot study
Vinod Prabhu1, Sadanand Prasadi1, Vishrabdha Pawar2, Aslam Shivani1, Alka Gore3
1 Department of Surgery, Bharati Vidyapeeth University, Sangli, Maharashtra, India
2 Department of Pathology, Bharati Vidyapeeth University, Sangli, Maharashtra, India
3 Department of P&SM Bharati Vidyapeeth University, Sangli, Maharashtra, India
|Date of Web Publication||12-Sep-2014|
Shreyas, Behind Central Warehouse, Miraj - 416 410, Maharashtra
Source of Support: None, Conflict of Interest: None
Objectives: An endeavor to look for cost effective method for dressing wounds to minimize loss of working hours. Three percent citric acid solution (CA) was used for dressing acute lower limb ulcers with the object of pH modulation of wounds at an early stage and to evaluate its effects on wound healing. This solution was compared with Edinburgh University solution of lime (Eusol) in a randomized double-blinded study. Appearance of healthy granulation was the end point of the study. Materials and Methods: An unicentric randomized double blinded study with a parallel design was used to compare patients treated with 3% CA and Eusol solutions, respectively. The results were analyzed using Chi-square and P value using Microsoft Excel and SPSS 22. Patients having fever, altered consciousness, high initial random blood sugar >350 mg%, positive urinary ketones, hypoproteinemia, signs of septicemia and vascular diseases were excluded. Results: Twenty patients were known diabetic out of which 11 were in the CA group, and the rest were in the Eusol group. It was observed that the average total stay in diabetic patients was 16.36 days in CA group and 31 days in Eusol group whereas it was 15.78 and 24.75 days respectively in the nondiabetic group. The ulcer granulation interval showed that the mean stay in CA group was 10.56 days while it was 20.04 days in the Eusol group. The cost of stay was less in CA group. Conclusions: It is concluded that 3% CA solution forms a good alternative for wound dressings that acts by modulating the wound pH to acidic levels thereby contributing to wound healing by increased fibroblast proliferation and probably increasing local oxygen concentration and reducing microbial growth and virulence.
Keywords: Acute ulcer, Edinburgh University solution of lime, granulation, wound healing, 3% citric acid solution
|How to cite this article:|
Prabhu V, Prasadi S, Pawar V, Shivani A, Gore A. Does wound pH modulation with 3% citric acid solution dressing help in wound healing: A pilot study
. Saudi Surg J 2014;2:38-46
|How to cite this URL:|
Prabhu V, Prasadi S, Pawar V, Shivani A, Gore A. Does wound pH modulation with 3% citric acid solution dressing help in wound healing: A pilot study
. Saudi Surg J [serial online] 2014 [cited 2019 Jan 16];2:38-46. Available from: http://www.saudisurgj.org/text.asp?2014/2/2/38/140690
| Introduction|| |
Wounds are classified into acute and chronic wounds with the addition of hard to heal wounds as a distinct entity.
Wound healing is a complicated and dynamic process that restores cellular structures and helps tissue healing. The wound healing process can be divided into three distinct phases viz. the inflammatory phase, the proliferative phase and the remodeling phase.  In these three broad phases, a complex series of events occur that includes chemotaxis, phagocytosis, neocollagenesis, collagen degradation, and collagen remodeling. In addition, angiogenesis, epithelization, and the production of new glycosaminoglycan's and proteoglycans are vital to wound healing. The end result of these biological processes results in the replacement of normal skin structures with fibroblast rich scar tissue. An acute wound will heal normally in uncompromised conditions. 
The most important requisite is to dress the wound with appropriate materials in order to allow the following:
(1) To keep the wound clean and free of infection (2) to reduce or eliminate pain and (3) to replace the missing tissue as much as possible.
The ideal wound dressing should be capable of wound protection, help in separation of devitalized tissue, promote fibroblast proliferation and help in the formation of healthy granulating surface to facilitate grafting as early as possible.
As many studies have been undertaken in chronic wounds, this study concentrates on acute wounds, as rapid healing response would obviate a lot of problems for the rural patients and avoid going into chronicity besides reducing the cost of treatment and minimizing loss of working hours. This study has been done in a rural medical college.
Many chemicals like hydrogels, hydrocolloids, betadine, Edinburgh University solution of lime (Eusol), honey, phenytoin, citric acid (CA) containing dressings have been used for application to wounds. ,
The aim of this study was to use 3% CA solution for application to wounds and see for changes that occurred in the wounds and the time taken for development of healthy granulation. Use of CA solution was preferred as it is easy to prepare, has minimal side effects with long shelf life and produces an acidic environment  as compared to Eusol which is a highly basic solution and inhibits fibroblasts. The purpose of this study was to test the efficacy of CA solution over Eusol by pH modulation. Lower limb, acute ulcers were preferred as we subserve predominantly rural farmer population that is more often susceptible to lower limb trauma.
| Materials and Methods|| |
This study is a unicentric randomized double blinded study conducted in rural medical college. As a tertiary referral center subserving a predominantly rural population, lower limb cellulitis, mainly due to diabetes and trauma constitute a large part of indoor patients that affects the economy and loss of working hours for most of the patients, thus affecting their lives. We planned to look for options other than conventional ones to minimize hospital stay and reduce the economic burden on the family.
Consent from Institutional Ethical Committee was obtained - bvdumc and h/sangli/iec/34/11dt20-12-11. As it was a pilot study the sample size was calculated using a statistical formula of α =5% and power = 90%; yielding a sample size of 25 in each group.
A parallel study design was used. The participants included all indoor patients of age 20-80 years with lower limb muscle deep ulcers <15 × 15 cm size and more than 10 days of history of trauma. Hemoglobin (Hb) >10 g%, nonhypoproteinemic (total proteins >6 g, albumin >3 g%, A/G ratio >1), glycosylated Hb <7 and vascular uncompromised participants were included.
Patients having fever, altered consciousness, high initial random blood sugar >350 mg%, positive urinary ketones, hypoproteinemia, signs of septicemia, vascular diseases, were excluded. The inclusion and exclusion factors were used to minimize confounding factors.
The flowchart showing the procedure followed is reproduced [Figure 1]. Objective of the trial was to assess the efficacy of 3% CA solution in healing acute lower limb ulcers comparing it with Eusol. Healing was assessed by reduction in colony count on culture, pathological evidence of granulation on biopsy, reduction of odor and pain using the visual analogue scale for pain (VAS) [Figure 2], range of limb movement using the manual muscle testing grading system (MMT) [Figure 3].
Development of healthy granulation was the end point of study. The ulcer-granulation (U-G) stay and date of discharge was recorded. The specific objective was to evaluate the speed of healing, hospital stay and cost effectiveness in the two groups. After informed consent from participants, the patients were allotted a computerized random number by a nonmedical person. After basic hematological tests, culture and edge biopsy were taken. Broad spectrum antibiotic (cefotaxime) was given initially awaiting culture reports and then changed or discontinued as per report. Associated comorbid factors such as hypertension, diabetes, IHD were treated accordingly for all participants.
About 3% analytical grade CA powder (Manufactured by Molychem, India) was dissolved in 100 ml sterile distilled water to obtain 3% CA solution. Similarly, Eusol was prepared by first dissolving 12.5 g of powdered chlorinated lime in about 100 ml of distilled water to form a paste and then add 12.5 g of boric acid powder to form an amalgamated paste and then add distilled water to make a 1 L solution. Allow the solution to stand and then filter it. Use the filtered solution as Eusol. These solutions were prepared by Biochemistry Department. Two similar sized bottles were used and labelled bottle 1 and bottle 2, one containing CA solution and the other containing Eusol.
Biopsy reports of the ulcer edge taken on 1 st and 7 th day were examined. As quantification of fibroblasts is difficult, we relied on the amount of granulation tissue that is, fibroblasts and neovascularization in 15 hpf with granulation seen in <7 hpf as sparse and above 7 hpf fields as satisfactory. The 7 th day biopsy was used to compare both arms.
The wound size was measured by ruler method or clock-face method depending upon the shape of the wound, and the pH was assessed by pH strips (Manufactured by Loba chem., India) starting day one and thereafter every 5 th day. The culture report indicated the colony count on all occasions except in mixed infections.
Pads soaked with a solution from either bottle 1 or 2 were used to dress the wounds in patients. The wounds were dressed daily once. On the day of admission, 7 th -day culture and edge biopsy were done. The wounds were assessed daily by two independent observers for pain, degree of slough separation, movements of a limb and hence observer error minimized.
The care provider (intern) dressed the wound from one of the two identical bottles, one containing 3% CA and the other containing Eusol, after cleaning the exudate on the wound surface with dry gauze. A maximum amount of 25 ml was used to soak the wounds for both solutions. If bottle no 1 was used for a particular patient then, the same bottled solution would be used for the subsequent dressing in the same patient till the final outcome is recorded. The bottle number was recorded for each patient so that at the final outcome the observer will record which bottled solution was used for a particular randomized number without being aware of what solution that bottle contained. The bottled solutions were obtained every week from the biochemistry department, which was not involved in the study beyond providing solutions. Personals (interns) dressing the wound were rotated every month.
It is appropriate to mention here that the participants, care providers and observers of outcome were blinded in this study.
At the end of the study, findings were compiled and grouped into those treated with bottle 1 and others treated with bottle 2 solution. Only after the inferences were drawn that the contents of the bottle were revealed.
The CA and Eusol solutions were then tested for any decline in pH over a period of 16 weeks using pH meter.
Statistical analysis was performed using mean, standard deviation, frequency, percentage as descriptive statistics. Unpaired t-test was used to check the significant difference between means of U-G days, total stay and cost comparison. Karl Pearson's correlation coefficient was applied to check the relationship between U-G days and cost for CA and Eusol groups. Statistical analysis was done using SPSS 22, IBM, Bangalore.
| Observation|| |
This study included 50 patients (n = 50) with 25 patients in each group that is, bottle 1 and bottle 2. Bottle 1 contained CA whereas bottle 2 contained Eusol.
The maximum number of patients were seen in 50-70 age group as these are the age groups that are frequently seen working outdoors in farms. There were only three female patients in this study given the predominance of male persons doing farm work.
There is no statistically significant difference in mean ages of patients where CA or Eusol were used (t = 0.613; P = 0.543).
The VAS and MMT was used to assess the severity of pain and evaluate the range of motions revealed no significant difference in both groups. The VAS score was in the range of 0-4 and the MMT score ranged from 3 to 5. There was no significant difference in the scores pre- and post-treatment in either groups.
The pH of wounds was monitored using pH strips which showed a pH ranging from 4 to 5 in the CA group, while the Eusol group showed a pH of 7-7.5, indicating that the wound environment was acidic in CA group and alkaline in Eusol group. None of the patients of both the groups complained of any local complaints such as skin irritation, burning sensation or pain aggravation on application.
Twenty patients were known diabetic out of which 11 were in the CA group, and the rest of the patients were in the Eusol group. It was observed that the average total stay in diabetic patients was 16.36 days in CA group and 31 days in Eusol group which was significant. Even in nondiabetic patients the average total stay was 15.78 days in CA group as compared to 24.75 in Eusol group. This is also observed in the U-G stay in both groups [Table 1]. That explains that the irrespective of diabetic or nondiabetic status the mean stay is less in the CA arm than the Eusol arm (P < 0.05). Diabetes as a comorbid factor was taken into account as it is a significant contributing factor in wound healing.
|Table 1: Comparison of the duration of stay in diabetic and non diabetic groups |
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The culture and sensitivity results in both groups indicate that the organisms were sensitive to antibiotics in six cases, resistant in seven and sterile in 12 cases in the CA group whereas the comparable numbers were 6, 9 and 10 in the Eusol group. The U-G days showed a significant difference in the CA group and the Eusol group with the stay being less in the former, in the resistant and sterile groups. However in culture sensitive cases there was no significant difference in stay probably due to the influence of specific antibiotics used [Table 2]. This is a significant finding that irrespective of the culture status the stay was less significantly in CA arm.
The ulcer granulation interval which indicated the speed of healing shows that the mean stay in CA group was 10.56 days while it was 20.04 days in the Eusol group [Table 3] indicating that the CA arm had a significantly reduced stay.
|Table 3: Comparison and statistical significance of Ulcer-Granulation (U-G) interval |
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There was no statistically significant reduction in size of the ulcers in both groups [Table 4]. The cost of the stay for the patients in CA group was 3396 (in rupee terms, calculated at 300/day) while it was almost double for the Eusol group [Table 5].
|Table 5: Cost comparison at 300/ - per day for U - G interval. (In rupee terms)|
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The histopathological examination on 7 th day showed an increase in fibroblasts and granulation tissue in the CA group when compared to the Eusol group.
The pH of the two solutions were further evaluated over a period of sixteen weeks which showed a fall in pH of Eusol solution from 9 to 7 whereas the pH of CA solution remained stable [Figure 4].
| Discussion|| |
Treatment of wounds has been a major challenge worldwide as it has a significant bearing on the socioeconomic condition of the society at large more so in developing countries. The very fact that numerous attempts have been made to identify a treatment system for wounds indicates the complexity of the problem. Our study has tried to evaluate CA dressing as a cost effective method for treating acute ulcers locally so that the patients can resume function as early as possible. CA is used as a preservative in many edible food items for its antimicrobial activity and is a weak organic acid, hence not harmful to the body. Within the European Union, it is referred to as E330. It is used as a pH adjusting agent in creams and gels, hair shampoo and as one of the active ingredients in the production of antiviral tissues. Its use in face packs for skin regeneration properties influenced us to use this as a wound dressing solution.
Citric acid has been in use for a very long time, first in dental care ,, and then subsequently in surgical practice. Authors worldwide have used hydrocolloid dressings,  pH altering solutions,  vacuum assisted closure,  nitrogen oxide releasing wound dressings to hasten the process of wound healing. For a substantial period of time Eusol  was used as a dressing solution in developing countries without studying the histological and pathological effects it had on wound healing. Our rural medical college hospital is also using Eusol. However, the volume of acute traumatic ulcer patients presenting late to the tertiary care center [Figure 5] leading to delay in wound healing and loss of working hours prompted us to look for an effective solution for dressing which would have a longer shelf life and be stable at room temperature which is fulfilled by CA solution. 
The use of CA was prompted by the concept of pH modulation.  Most of the pathogens thrive in a pH of more than 6. The more the pH is on the acidic side the chances of organisms surviving are bleak. Monitoring of pH is a challenge as it is difficult to measure the pH of the milieu interior of the ulcer and hence surface pH is monitored, after cleaning the exudate. By trying to change the wound pH using CA, we attempted to increase the oxygen concentration in the wounds thereby facilitating the migration and proliferation of fibroblasts and inhibit bacterial growth , which is evident from the pathological and microbiological evidence, that showed an increase in fibroblast concentration in the CA group whereas in the Eusol group the high alkaline pH inhibits fibroblast migration.  Lowering of pH by 0.6 units increases release of oxygen by almost 50%. Due to limited access to advanced instrumentation we used pH strips which gave a fairly good pH value. This concept of pH strip has been attempted to be incorporated in certain wound dressings to indicate the pH status of wounds.  The pH of wounds in our series indicate an acidic pH during the healing phase which gradually increased toward alkaline pH as granulation increased, thus proving the significance of pH in wound healing.  The pH strip method used in this study indicated a pH range of 4-5 in CA treated group whereas it was 7-7.5 in the Eusol group thereby implying that CA provides a conducive acidic environment that aids in wound healing. This same principle is used in the treatment of wounds with honey which apart from its antimicrobial properties produces an acidic environment thus facilitating wound healing.  Besides, a moist dressing aids in slough separation and prevents tissue dehydration. 
Diabetes is a significant contributor in delaying wound healing. However, we found that a statistically significant reduced stay in CA treated group as compared to the Eusol group irrespective of diabetic status.
The culture and sensitivity reports taken on day 1 and day 7 also revealed that the colony counts of patients treated with CA reduced in some cases, whereas there was no change in the Eusol group. However in many cases, there was a mixed bacterial infection hence the reduction of colony count could not be accurately documented in each case. The organisms commonly isolated were pseudomonas auregenosa and proteus group. As is well known these organisms are very virulent and resistant to routine antibiotics, but with CA dressings, they did not prove to be an impediment to healing. It was found that in all groups of patients that is, sensitive, resistant and sterile groups the U-G days were significantly less as compared to the Eusol group, but a statistical significance was seen in resistant and sterile groups only as specific antibiotics used in the sensitive group could have had influence on healing.
Similarly, measurement of wound size is challenging. Ideally if facilities are available stereophotogrammetry measurement of size is most reliable, however we used the clock face and ruler method to measure the size ,, which the authors submit is a crude method and hence, although we have recorded a reduction in wound size in the group treated with CA it has no statistical significance although two observers have independently evaluated the wounds and confirmed the size reduction thereby minimizing observer error.
The histopathological examination showed that fibroblasts were seen in substantial concentration in CA treated cases on the seventh day of admission [Figure 6]a with fibroblasts and vascularization seen in more than 12 hpf thereby implying that the fibroblast migration is not hampered in this group when compared to the Eusol group where scarce fibroblast concentration [Figure 6]b < 7 hpf suggested the deleterious effect of an alkaline pH on wound healing.
The cost of treatment in the CA group was considerably less when compared to the other group which is significant, considering that this institute is a rural setup in a geography that is marred by increased number of farmer deaths due to socio economic reasons.
|Figure 6: (a) H and E, ×10 showing substantial granulation tissue in citric acid treated cases (b) H and E, ×10 showing sparse granulation in Edinburgh University Solution of Lime treated cases|
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Although wounds are classified as acute and chronic wounds, it appears that a definitive subset of wounds called as hard to heal wounds do exist, that are resistant to healing due to various factors. Although the wounds in this study do not fit into this group in the strictest sense many of factors that are responsible for hard to heal, wounds exist in this study that are likely to delay healing viz. aging population, size of wounds, social factors, delay in healthcare access, etc. 
Recognition of five factors that are predictive of delayed wound healing are of prime importance in order to intervene, to facilitate smooth healing, obviating the long stay and inflated costs of treatment. ,, These five factors are wound location, wound infection, wound size, wound duration and patients age. Another acronym for wounds is aptly summarized in the word DOMINATE  which outlines precautions to be taken in managing wounds.
All the above factors incline toward a wound pH that is conducive to healing and a successful skin graft acceptance. ,
Further, in order to test for stability of pH of the two solutions at room temperature we tested the pH of these solutions over a period of 16 weeks and found that the pH of the CA solution was 2.0 when compared to the Eusol solution whose baseline pH was 9 which came down to 7 for the comparable period. These solutions were tested for pH every week from the same bottles every time.
However, the authors concede that as it is a pilot study the sample size is small to come to any definitive, unambiguous conclusions, but still the results derived are encouraging and could be extrapolated to be used in a larger sample size with minimum exclusion criteria.
| Conclusion|| |
This study concludes that CA modulates the wound pH to acidic so as to facilitate rapid wound healing thereby reducing the cost of treatment and preventing the loss of working hours there by minimizing economic loss. The study will give an impetus for use of CA solution in larger ulcers with minimum exclusion criteria.
| Acknowledgement|| |
This study would not have been possible without the help of Departments of Pathology, Microbiology and Biochemistry. The authors also wish to thank the interns and the nursing staff who wholeheartedly supported this study and last, but not the least our institute for providing all the logistics. The authors also recognize the valuable inputs from Dr. B. S. Nagoba, Head of Research MIMS and Dr. Raghvendra Kulkarni, Head of the Department of Microbiology, SDM Medical college, Dharwad. We thank all others who could not be mentioned for obvious reasons.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6]
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5]