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\begin{document}


\def\RunningAuthor{Sheela Kumar Gujjari et al.}
\firstPage{2530}
\articleType{Original Article}
\receivedDate{15 Aug 2021}
\acceptedDate{20 Sep 2021}
\revisedDate{18 Sep 2021}
\journalVolume{2021, 12}
\journalIssue{4}
\journalDoi{ijrps.v12i4.4901}
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\title{\textbf{To Evaluate Aloe Vera- Chitosan Based Surgical Dressing to Chitosan Dressing Alone on the Healing of Palatal Donor Sites - A Pilot Study}}
\author{Archana~R~Sankar\textsuperscript{1},  
            Sheela~Kumar~Gujjari\textsuperscript{*}\textsuperscript{1}, 
            Kulkarni~P~K\textsuperscript{2}~\\[5pt]{\textsuperscript{1}Department of Periodontology, JSS Dental College and Hospital, Mysuru, Karnataka, India}~\\{\textsuperscript{2}Department of Pharmaceutics, JSS College of Pharmacy, Mysuru, Karnataka, India}}

\begin{abstract}
The present study was undertaken to clinically assess the synergistic effect of topically applied Aloe vera Chitosan to Chitosan alone on the healing of palatal donor sites in free gingival graft surgical procedures. 20 subjects (10 per group) were enrolled into this triple blinded randomized clinical trial, designed to evaluate the efficacy of Aloe vera Chitosan to Chitosan on free gingival graft surgical wounds. Subjects were assigned to either Group A (aloe vera + Chitosan) or Group B (Chitosan alone). Visual Analog Scale(VAS) and Wound healing index were recorded at 7\ensuremath{^{th}} day, 14\ensuremath{^{th}} day and 21 days postoperative. Results showed no statistical significance when considering VAS. However there was a statistical significance in wound healing (p=0.04) in group A when compared to group B from 14- 21\ensuremath{^{st}} day postoperative. It is the first human trial and was a pilot study to assess the synergistic effect of Aloe vera and Chitosan on wound healing. The study has shown that a combination of both acts synergistically in accelerating healing especially in areas where faster healing is necessary to provide overall comfort to the patient. It is therefore a futuristic and a promising material as a surgical dressing with further longitudinal trials.
\end{abstract}\def\keywordstitle{Keywords}

\begin{keywords}Aloe vera,\newline Chitosan,\newline Free Gingival Graft,\newline Gingival Recession,\newline Healing,\newline Visual Analog Scale
\end{keywords}
\twocolumn[ \maketitle {\printKwdAbsBox}]
\makeatletter{\raggedright\textsuperscript{*}Corresponding Author\par Name:\ Sheela~Kumar~Gujjari~\\ Phone:\ 9448836777~\\  Email:\ sheelagujjari@gmail.com
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\section{Introduction}
Wound healing is a complex and dynamic process where cellular structures and tissue layers are formed which results in function and restoration of injured tissue. A coordinated effort of epithelial cells with several other cell types including fibroblasts, endothelial cells, macrophages and platelets are noted \unskip~\citep{1225367:23301557,1225367:23301558}. The migration, infiltration, proliferation and differentiation of these cells culminates in an inflammatory response, the formation of new tissue and ultimately closure of the wound. The oral cavity provides a unique environmental challenge for the epithelial healing of oral wounds produced during various periodontal procedures such as mucogingival surgeries where graft harvesting is required from donor site. The ability of the oral mucosa to repair itself after wounding is remarkable as it involves large exposure of the surgical area leading to pain, discomfort and healing by secondary intention.

Various dressings have been used in periodontal therapy to protect the wound, the most popular being Eugenol (Wondr Pack) and Non Eugenol(Coe Pak) \unskip~\citep{1225367:23301557}. Whether to place a pack or not is still a controversy \unskip~\citep{1225367:23301557} but in certain conditions where there is exposure of large wound area such as in free gingival grafts, gingivectomies, depigmentation, there is still a need to protect the wound. Apart from these dressings ozonated oil \unskip~\citep{1225367:23301558}, erythropoietin \unskip~\citep{1225367:23302042}  and plant extracts have also been tried to protect the wound in the palatal donor site which lead to overall patient comfort and stabilization of clot which is an essential element for wound healing. Though various dressings have shown improvement, the search is still on for an ideal surgical dressing material.

Various animal studies \unskip~\citep{1225367:23302095,1225367:23302099}  have demonstrated the healing potential of Aloe vera as it increases the blood supply, oxygenation, cell proliferating activity \unskip~\citep{1225367:23302133,1225367:23302140}, antibacterial, antiviral, antifungal and anti inflammatory properties \unskip~\citep{1225367:23302141,1225367:23302142,1225367:23302144}  which has kindled an interest among researchers in its usage as a dressing. Recently much of interest has also been developed towards Chitosan, a biopolymer obtained from the deacetylation of chitin a naturally occurring and abundantly available (in marine crustaceans) biocompatible polysaccharide \unskip~\citep{1225367:23302145}. 

Evidence from animal studies have shown excellent biocompatibility, biodegradability, non-toxicity and non-allergenic nature and its potential role as an antimicrobial and anti inflammatory substance, ultimately resulting in favourable wound healing \unskip~\citep{1225367:23302148,1225367:23302231,1225367:23302183}. It has been hypothesized that the mechanism of action of Aloe vera and Chitosan on wound healing may be connected in part to their antimicrobial effect but also with its ability to promote the liberation of growth factors, activate local antioxidant mechanisms and promote tissue repair. 

Chitosan is shown to be highly mucoadhesive \unskip~\citep{1225367:23302312} which helps in the retention of the dressing in the palate. An attempt was made to incorporate Aloe vera into Chitosan as Chitosan is essential to stabilize the polysaccharides present in Aloe vera which in turn helps to keep its natural biological activity stable \unskip~\citep{1225367:23302360}. 

Although abundant animal studies are done with Aloe vera and Chitosan in the field of medicine, there are no human studies with a combination of both. Thus, this study was done to evaluate the synergistic efficacy of Aloe vera Chitosan based surgical dressing to Chitosan surgical dressing alone on healing of palatal surgical wounds.
    
\section{Materials and Methods}
\textbf{Materials used in the study}


  
  \begin{enumerate}
  \item \relax Commercially obtained high molecular weight Chitosan, with a degree of deacetylation above 85\% was purchased from Everest Biotech, Bengaluru, Karnataka.
  \item \relax Commercially available Aloe vera gel (Patanjali) obtained from local markets.
  \end{enumerate}
  \textbf{Preparation of Chitosan{\textendash}Aloe Vera Blend \ensuremath{^{}}}

The gel form of chitosan was obtained by adding 5 g of commercially purchased chitosan to 100 mL of water in a blender for 10 min until the hydrogel was formed which was then packed in a sterile tubes, sealed and UV sterilized later. Preparation of Aloe vera Chitosan comprised of adding commercially obtained Aloe vera gel into the Chitosan gel (at a ratio of 1:2 v/v) which wasstirred thoroughly to obtain a uniform mixture and was dispensed into sterile aluminium tubes \unskip~\citep{1225367:23302360,1225367:23302362}. Both gels had the same texture and colour. The tubes were coded as A and B. The coding was done by a Professor in Pharmacy which was decoded after the study (Figure~\ref{f-819fa04b18ab}).


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\caption{\boldmath {CONSORT chart}}
\label{f-88426db37209}
\end{chart*}
\egroup

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\caption{\boldmath {VAS}}
\label{f-7d92c86c1777}
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\egroup

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\caption{\boldmath {Healing Index Scores}}
\label{f-53b3d1977b29}
\end{graph*}
\egroup

\bgroup
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\makeatother 
\caption{\boldmath {Preparation of ablend of Aloe vera and Chitosan. (a \& b) addition of Chitosan to Aloevera gel \& mixing both (c \& d) transferring the mix to sterile packing tubes and sealing the ends}}
\label{f-819fa04b18ab}
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\egroup

\bgroup
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\makeatother 
\caption{\boldmath {Root Coverage Using Free Gingival Graft. (a) donor site for graft harvesting. (b \& c) graft placed onto recipient bed and sutured (d) application of the gel either Aloevera - Chitosan or Chitosan alone}}
\label{f-81b7ad9ae1bf}
\end{figure*}
\egroup

\bgroup
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\centering \makeatletter\IfFileExists{images/3eb4c535-60e9-4d83-814a-fcc21fdb091a-upicture6.png}{\includegraphics{images/3eb4c535-60e9-4d83-814a-fcc21fdb091a-upicture6.png}}{}
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\caption{\boldmath {Aloe Vera and Chitosan Group}}
\label{f-04b5b0bd92d2}
\end{figure*}
\egroup

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\caption{\boldmath {Chitosan Group}}
\label{f-2e31d19b1765}
\end{figure*}
\egroup
The patients were selected from the outpatient's department of Periodontology, JSS Dental College and Hospital, Mysuru. By purposive sampling, patients of both sexes who fulfilled the inclusion criteria and who had given the informed consent were enrolled for this study. Randomization was done by lottery method and the patients were allocated into two groups (A) and (B) (Chart~\ref{f-88426db37209}).

Group A- `` Aloe vera- Chitosan dressing''

Group B-`` Chitosan dressing alone''

The inclusion criteria comprised of systemically healthy patients within the age group of 20-40 both males and females, patients willing to undergo soft tissue grafts, Teeth showing localized recession in upper or lower anterior sextant, Subjects with Millers class I/II \ensuremath{\geq }3mm who required soft tissue grafting, who were able to and willing to follow study procedure and instructions.

The exclusion criteria comprised of patients on medications such as antibiotics, corticosteroids, hormonal drugs in the past 2 months\textbf{, } systemic conditions known to affect the outcomes of periodontal therapy immunocompromised patients\textbf{, } smokers\textbf{, } pregnant and lactating women\textbf{,} subjects with prominent root surface\textbf{, } soft tissue loss interproximally, periodontal pockets\textbf{, } grade II or higher mobility\textbf{, } cervical abrasions restored.

\textbf{Sample Size Calculation}

The sample size was estimated based on hypothesis testing for two means(equal variance assumed) using n Masters software. The sample size was estimated to be 8 per group at a mean difference of 0.9, effect size 1.38, 80\% power and 5\% level of significance. Finally the sample size was rounded off to 10 per group (20 patients) anticipating some degree of drop out.

\textbf{Study Design}

The study was a parallel arm, triple blinded, randomized controlled study which was carried out from August 2019 to February 2020. By purposive sampling, 10 subjects per group were selected. The randomization process was performed by an examiner blinded to the study. The allocation was concealed within opaque envelopes with numbers on it until immediately before surgery to determine which subjects would fall into Group A and Group B.

\textbf{Ethical Clearance and Informed Consent}

Institutional Review Board (IRB) of JSS Academy of Higher Education and Research approved the study protocol. A prior written informed consent was taken from the participants. All principles outlined in the Declaration of Helsinki(1964, revised in 2008) on experimentation involving humans were observed.

\textbf{Pre Surgical Procedure}

Patients fulfilling the inclusion criteria and have willingly given informed consent underwent routine hematological investigations and phase I~therapy comprising of scaling along with root planing.

\textbf{Clinical Data Collection}

Clinical parameters were recorded at baseline and on recall visits at 7\ensuremath{^{th}}, 14\ensuremath{^{th}}, 21\ensuremath{^{st}} days and 1 month. The clinical parameters recorded were


  
  \begin{enumerate}
  \item \relax Wound Healing Index of Landry, Turnbull and Howley \unskip~\citep{1225367:23302366}.
  \item \relax Wong Becker Visual analog scale
  \end{enumerate}
  After examination and completion of initial periodontal therapy subjects with full mouth plaque score (FMPS){\textless}20\% were selected for surgery.  Before surgery, an impression of the palate was made to prepare a stent for donor site protection. All surgeries were performed by the same surgeon blinded to the study.  The surgery was carried out according to the method of Sullivan and Atkins \unskip~\citep{1225367:23302371}.

After asepsis, the sites (donor+recipient) were anesthetized using 2\% lidocaine with 1:80,000 adrenaline. The recipient bed was prepared by de epithelialization. A standardized 10 \ensuremath{\times} 15 mm template was used to obtain an identical palatal wound area. The donor site for graft harvesting was chosen to be distal to canine (to avoid the rugae) upto the 1\ensuremath{^{st}} molar palatally \unskip~\citep{1225367:23302426}. Harvesting grafts from the same site in all patients resulted in a uniform technique to compensate for the differences in healing potentials. A rectangular shaped graft of sufficient thickness (upto 1.5mm) was harvested by sharp split thickness dissection. Care was taken to place the incision at least 2 mm apical from the gingival margins.

After graft removal wet gauze was compressed to the donor site with finger pressureuntil hemostasis was achieved. The graft was then sutured to recipient site using 5-0 vicryl. After suturing of graft, Group A received Aloe vera with Chitosan dressing in the donor site and Group B received only Chitosan dressing which was done using sterile applicator tips and before the placement of stent (Figure~\ref{f-81b7ad9ae1bf}). A soft stent was used to protect the wounded sites in both groups. Patients were advised to keep the stent in place for 1 week after which it was carefully removed for evaluation.

\textbf{Postoperative Care}

Participants were instructed to avoid eating and drinking acidic/hot food. Manifestations of any allergic reaction like irritation, itching, etc. were observed for 1-hour post-surgery, subsequent to which patients were discharged along with postoperative instructions. The patients were instructed to maintain their oral hygiene with regular home care, except in the operated area. Antibiotic (Amoxicillin 500mg) was prescribed for 5 days and Analgesic (Aceclofenac 100 mg) was advised to be taken only if it was necessary on a 'need to treat' basis. The patients were also asked to make a note of the number of analgesics consumed along with the duration in a notebook.

At each appointment on the 7\ensuremath{^{th}}, 14 and 21\ensuremath{^{st}} days postoperatively, Visual Analog Scale ranging from 0 (no pain) to 10 (worst imaginable pain) \unskip~\citep{1225367:23302446}  and wound healing were assessed. Wound healing was assessed clinically using a Healing index by Landry et al., which grades the wound on a scale of 1{\textendash}5 (Table~\ref{tw-0e276916aba2}) where 1 indicates very poor healing and 5 indicates excellent healing \unskip~\citep{1225367:23302366,1225367:23302447}. The scores were given by a different examiner. The patients were also followed up routinely for a period of 3 months.


\begin{table*}[!htbp]
\caption{\boldmath {Healing Index} }
\label{tw-0e276916aba2}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.3543000000000001\linewidth-2\tabcolsep}p{\dimexpr.6457\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack Healing Index Score & Clinical Findings\\
\tblmidrule 
\cAlignHack Very poor (1) &
  Tissue color: \ensuremath{\geq }50\% of gingiva red \mbox{}\protect\newline Response to palpation: Bleeding \mbox{}\protect\newline Granulation tissue: Present \mbox{}\protect\newline Incision margin: Not epithelialized, with loss of epithelium beyond incision margin \mbox{}\protect\newline Suppuration: Present\\
\cAlignHack Poor (2) &
  Tissue color: \ensuremath{\geq }50\% of gingiva red \mbox{}\protect\newline Response to palpation: Bleeding \mbox{}\protect\newline Granulation tissue: Present \mbox{}\protect\newline Incision margin: Not epithelialized, with connective tissue exposed\\
\cAlignHack Good (3) &
  Tissue colour: \ensuremath{\geq }25\% and{\textless}50\% of gingiva red \mbox{}\protect\newline Response to palpation: No bleeding \mbox{}\protect\newline Granulation tissue: None \mbox{}\protect\newline Incision margin: No connective tissue exposed\\
\cAlignHack Very good (4) &
  Tissue colour: {\textless}25\% of gingiva red \mbox{}\protect\newline Response to palpation: No bleeding \mbox{}\protect\newline Granulation tissue: None \mbox{}\protect\newline Incision margin: No connective tissue exposed\\
\cAlignHack Excellent (5) &
  Tissue color: All tissues pink \mbox{}\protect\newline Response to palpation: No bleeding \mbox{}\protect\newline Granulation tissue: None \mbox{}\protect\newline Incision margin: No connective tissue exposed\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Demographic Data} }
\label{tw-01fcf02d6b9f}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.1708\linewidth-2\tabcolsep}p{\dimexpr.11430000000000003\linewidth-2\tabcolsep}p{\dimexpr.12849999999999998\linewidth-2\tabcolsep}p{\dimexpr.1708\linewidth-2\tabcolsep}p{\dimexpr.12329999999999998\linewidth-2\tabcolsep}p{\dimexpr.1323\linewidth-2\tabcolsep}p{\dimexpr.16\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack  & \cAlignHack N & \cAlignHack Mean & \cAlignHack Std. Deviation & \cAlignHack Std. Error & \cAlignHack Minimum & \cAlignHack Maximum\\
\tblmidrule 
\cAlignHack M &
  \cAlignHack 15 &
  \cAlignHack 34.0000 &
  \cAlignHack 5.2768 &
  \cAlignHack  &
  \cAlignHack 32.00 &
  \cAlignHack 47.00\\
\cAlignHack F &
  \cAlignHack 5 &
  \cAlignHack 30.7500 &
  \cAlignHack 7.5240 &
  \cAlignHack 3.32603 &
  \cAlignHack 30.00 &
  \cAlignHack 44.00\\
\cAlignHack Total &
  \cAlignHack 20 &
  \cAlignHack 39.1444 &
  \cAlignHack 6.6682 &
  \cAlignHack 1.44706 &
  \cAlignHack 31.00 &
  \cAlignHack 47.00\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {VAS} }
\label{tw-7561e296a489}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.2298\linewidth-2\tabcolsep}p{\dimexpr.2523\linewidth-2\tabcolsep}p{\dimexpr.2477\linewidth-2\tabcolsep}p{\dimexpr.2702\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack VAS & \cAlignHack Group A & \cAlignHack Group B & \cAlignHack P Value\\
\tblmidrule 
\cAlignHack 7 Days &
  \cAlignHack 8.0 &
  \cAlignHack 8.5 &
  \cAlignHack \\
\cAlignHack 14 Days &
  \cAlignHack 4.9 &
  \cAlignHack 4.7 &
  \cAlignHack 0.07\\
\cAlignHack 21 Days &
  \cAlignHack 1.6 &
  \cAlignHack 1.4 &
  \cAlignHack \\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Intergroup Comparison of Healing Index Scores (Donor Site)} }
\label{tw-9345bbf47ad5}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.389\linewidth-2\tabcolsep}p{\dimexpr.19389999999999997\linewidth-2\tabcolsep}p{\dimexpr.18960000000000004\linewidth-2\tabcolsep}p{\dimexpr.2275\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack Postoperative day & \cAlignHack Group A & \cAlignHack Group B & \cAlignHack Significance\\
\tblmidrule 
\cAlignHack 7\ensuremath{^{th}} day postoperative &
  \cAlignHack 3.2\ensuremath{\pm}0.573 &
  \cAlignHack 2.8\ensuremath{\pm}0.675 &
  \cAlignHack 0.273\\
\cAlignHack 14\ensuremath{^{th}} day postoperative &
  \cAlignHack 4.2\ensuremath{\pm}0.667 &
  \cAlignHack 3.1\ensuremath{\pm}0.771 &
  \cAlignHack 0.089\\
\cAlignHack 21 days postoperative &
  \cAlignHack 5\ensuremath{\pm}0.1 &
  \cAlignHack 4.0\ensuremath{\pm}0.516 &
  \cAlignHack 0.004\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}
\textbf{Statistical Analysis}

The results were analyzed using descriptive statistics, one way ANOVA and paired t-test using SPSS version 22 software.
    
\section{RESULTS }
The study aimed to evaluate the synergistic efficacy of Aloe vera Chitosan based surgical dressing to Chitosan surgical dressing alone on healing of palatal surgical wounds. It was a parallel arm, triple blinded randomized controlled study wherein 10 subjects per group were selected and received either a combination of Aloe vera Chitosan or Chitosan alone.

After surgery the donor site of both groups received either a combination of Aloe vera- Chitosan or Chitosan dressing alone. No untoward incidents were reported. Patients were assessed for VAS and wound healing at 7\ensuremath{^{th}} day, 14\ensuremath{^{th}} day, 21\ensuremath{^{st}} day and 1 month postoperative as complete epithelilization requires atleast 1 month according to a study by \unskip~\cite{1225367:23302042}.

A total of 20 participants were selected for study, 15 of which were males and 5 were females. The mean age of males were 34.00 \ensuremath{\pm} 5.2. The mean age of females were 30 \ensuremath{\pm} 7.5 (Table~\ref{tw-01fcf02d6b9f}).

VAS showed reduction in pain from baseline to 21 days although no statistical significance was noted between groups (p=0.07)(Table~\ref{tw-7561e296a489}, Graph~\ref{f-7d92c86c1777}).

The mean healing scores as per the healing index in the donor site in group A was 3.2 at 7\ensuremath{^{th}} day postoperative and 5.0 at 21 days. In group B the mean healing scores were 2.8 on the 7\ensuremath{^{th}} day which improved to 4.0 at 21 days postoperative with a statistical significance of 0.004 favour of Group A (Table~\ref{tw-9345bbf47ad5}, Graph~\ref{f-53b3d1977b29}, Figure~\ref{f-04b5b0bd92d2} and Figure~\ref{f-2e31d19b1765}).
    
\section{DISCUSSION }
The study aimed to evaluate the synergistic efficacy of Aloe vera Chitosan based surgical dressing to Chitosan surgical dressing alone on healing of palatal surgical wounds. It was a parallel arm, triple blinded randomized controlled study wherein 10 subjects per group were selected and received either a combination of Aloe vera Chitosan or Chitosan alone.

Aloe vera and Chitosan were used synergistically because both have very good healing potential along with pain reduction and anti inflammatory property. In addition to this, Chitosan was used as it is highly mucoadhesive \unskip~\citep{1225367:23302312} which helps in the retention of the dressing in the palate. It has also been shown to stabilize the polysaccharides present in Aloe vera which in turn helps to keep its natural biological activity stable \unskip~\citep{1225367:23302360}.

The randomization into group A and B were performed by an examiner blinded to the study and a calibrated examiner recorded the clinical parameters. The primary outcome of the study was to assess pain and wound healing. Pain was assessed using Wong Becker Visual analog scale and healing was assessed by Landry et al., healing index at 7\ensuremath{^{th}} day, 14 day and 21\ensuremath{^{st}} day postoperative Selection of palatal donor site as a healing assessment model is based on permitting standard wound creation, and easy access for clinical observation by means of being an open and shallow soft tissue healing model.

The area between distal finish lines of canine and first molar tooth - one of the most suggested donor site was selected for graft harvesting due to its anatomic advantages with regard to the course of the greater palatine artery and also to be able to make a comparison with the results of previous studies evaluating donor site outcomes following FGG \unskip~\citep{1225367:23302449,1225367:23302605}. All the surgeries were performed by the same periodontist and a standardized template was used to obtain a uniform thickness of the wound. Analgesic(Aceclofenac 100mg) was prescribed postoperatively as a rescue medicine as pain peaks 24 hours postoperatively \unskip~\citep{1225367:23302654}. The patients were also advised to make a note of how many analgesics were consumed during the week.

All patients consumed analgesics for 3 days as and when needed after surgery. 5 patients continued for 4 days postoperatively after which analgesics were discontinued. Pain was assessed at various intervals at 7\ensuremath{^{th}}, 14\ensuremath{^{th}} and 21stday. Wong Becker's scale was used as it is easy to administer and requires no equipment except for the photocopied faces. The absence of smiles and tears in this faces enables the patient to understand the scale better \unskip~\citep{1225367:23302699}.

A reduction in VAS scores were seen in both groups from 7\ensuremath{^{th}} to 21\ensuremath{^{st}} day postoperative, but was not statistically significant between the groups and at various time intervals. Though studies have shown the analgesic effects of aloe vera and chitosan \unskip~\citep{1225367:23302776,1225367:23302777}, in this study a statistical significance could not be appreciated. This could be attributed to application of the gel only once in both the groups and washing away of the gels by saliva thereby lowering its analgesic effect.

The patients reported that they had taken analgesics as rescue medication. So the gels did not show any analgesic effect, though anti-inflammatory effect was observed.

Healing index showed a statistical significance of p=0.004 in favour of Group A(Aloe vera Chitosan) between 14\ensuremath{^{th}} to 21\ensuremath{^{st}} days. This is in accordance to various studies where Aloe vera and Chitosan were used individually and showed very good healing along with pain reduction which was contradictory to this study. Various other studies also showed Aloe vera gel to have cell proliferating-promoting activity and also stated that aloe vera gel improved wound healing by increasing blood supply, which increased oxygenation as a result \unskip~\citep{1225367:23303063}.\ensuremath{^{}}

Two clinical trials on wound healing in genital herpes was researched and was concluded that Aloe vera showed better healing compared to placebo at the end of 2 weeks \unskip~\citep{1225367:23303084}.

The possible mechanism of Aloe vera in wound healing was explained in a systematic review. It was attributed that wound healing was due to Glucomannan and plant growth hormone gibberellins which interacted with growth factor receptors of fibroblast and stimulated its activity and proliferation for the increase of collagen synthesis when applied topically and on oral administration \unskip~\citep{1225367:23303373}.

T. Reynolds in 1999 evaluated the healing benefits of aloe vera in fresh minor recurrent apthous ulcer and showed reduction in the ulcer size, erythema, exudation, pain score, wound size and wound healing period \unskip~\citep{1225367:23303063}. A double blinded, split mouth, randomized clinical trial assessing the healing of sites treated with fresh \textit{Aloe vera }extracts after periodontal flap surgery was conducted. Better healing was observed in test sites in comparison to control sites in the first postoperative week \unskip~\citep{1225367:23303063}, which is in accordance with our study. So better healing could be contributed by Aloe vera component which would have acted by mechanisms similar to that of the above mentioned studies.

It has been observed in another study that chitosan hydrogel was able to promote cell adhesion and proliferation making it a promising material for wound healing. Excellent pain relief was also conferred by Chitosan when it was applied as a topical agent to open wounds, such as burns, skin abrasions, skin ulcers and skin grafted areas \unskip~\citep{1225367:23302777}. Similarly Chitosan gel used in this study also could have shown similar effects. Thus, a combination of Chitosan and Aloe vera would have improved wound healing.

An in vitro study proved that addition of aloe vera to chitosan enhanced antimicrobial potential and exhibits spindle-like shape typical of fibroblasts, suggesting good adherence to the surface and enhancement of wound healing \unskip~\citep{1225367:23303376,1225367:23303379}  which would have lead to beneficial effects in terms of wound healing in this study.

From the results of the study, Aloe vera - Chitosan combination acted synergistically compared to Chitosan alone in healing of palatal donor sites. No statistical significance was noted when considering pain but a combination of aloe vera and chitosan was statistically significant when considering healing of palatal wounds, suggesting its role as a futuristic dressing material with further longitudinal trials.

\textbf{Limitations of the Study}

Cytotoxicity and biodegradability tests in this study were not performed although animal and in vitro studies have confirmed the same.

Antimicrobial properties were not assessed. The study did not assess stability, absorption and release patterns of the components in the gels. Wound healing markers such as Matrix metalloproteinases (MMPs) could have been assessed.

\textbf{Future Perspective }

Although abundant evidence is available on aloe vera and chitosan, there is still a need to manipulate the physical properties of identified components and systems which are challenging to achieve the ideal objectives of a surgical dressing.
    
\section{Conclusion}
This was a preliminary study done to assess the synergistic effect of aloe vera chitosan to chitosan alone as a surgical dressing in palatal donor sites. It is the first human trial where a combination of both were used in the oral cavity to assess pain and wound healing in the palate following harvesting of free gingival graft. The materials are economical and compatible to the patient. Although statistical significance was noted in aloe vera chitosan group in terms of wound healing further longitudinal studies with detailed characterization is essential to prove its potential as a wound dressing material.

\textbf{Funding Support}

The authors declare that they have no funding support for this study.

\textbf{Conflict of Interest}

The authors declare that there is no conflict of interest.
    

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