**1. Introduction**

Silymarin is a flavonolignan derived from milk thistle that contains silibinin, isosilibinin, silychristin, isosilychristin, and silydianin, as well as taxifolin. Amongst its various constituents, silybin with its two diastereoisomeric compounds, namely silybin A and silybin B, are present in a higher percentage (around 70%) and contribute to the biological effect exerted by silymarin [1]. Silymarin has been used as a therapeutic agent in an array of liver disorders such as chronic liver disorders, cirrhosis, hepatocellular carcinoma, alcohol abuse, non-alcoholic fatty liver disease, virus-related liver damage and end stages of different hepatopathies because of its inherent antiviral, antioxidant, anti-inflammatory, and antifibrotic

**Citation:** Venugopal, D.C.; Senthilnathan, R.D.; Maanvizhi, S.; Madhavan, Y.; Sankarapandian, S.; Ramshankar, V.; Kalachaveedu, M. Preparation and Characterization of Silymarin Gel: A Novel Topical Mucoadhesive Formulation for Potential Applicability in Oral Pathologies. *Gels* **2023**, *9*, 139. https://doi.org/10.3390/ gels9020139

Academic Editor: Shige Wang

Received: 31 December 2022 Revised: 27 January 2023 Accepted: 2 February 2023 Published: 7 February 2023

**Copyright:** © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

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**Preformulation studies** 

**Organoleptic characters**

**Evaluation parameter for gel** 

Organoleptic character

Drug release

Melting point 150 °C

pH 6.42

Spreadability 23.75

Theoretical Drug content 32.77 ± 0.20 mg/g

properties [2]. Silymarin has also demonstrated chemopreventive action and antimetastatic activity in *in vitro* and *in vivo* conditions in various cancers, chiefly gastrointestinal cancers and also in other cancers such as colorectal and pancreatic cancers [3]. Silymarin-mediated apoptosis, through increased cleavage of caspase 8, has been demonstrated using *in vitro* studies using oral cancer cell lines [4]. Silymarin has been found to have a good safety profile without toxicity, even after prolonged administration [5]. Although there has been a long history of silymarin use in various systemic forms such as tablets, capsules, and suspension, currently randomized controlled trials (RCT) using a topical formulation of silymarin have been employed as a therapeutic option.

Gels offer certain advantages over other formulation types, including quicker drug release, smoother delivery, and better bio-compatibility and mucoadhesivity. Gels permit adherence to the oral mucosa at the site of the lesion and rapid elimination by regular catabolic routes [6]. A topical gel formulation for skin, prepared by Sampatrao and the team showed drug content, pH, and spreadability of the formulation to be 96.6%, 6.8 and 25.45 gm.cm/s, respectively, with a maximum drug release of 96.30% throughout 3 h. The gel demonstrated pseudoplastic flow properties without showing any acute skin irritancy [7]. A randomized, double-blinded, placebo-controlled clinical trial evaluated the preventive effect of a silymarin 1% gel in comparison with a placebo, on the occurrence of radiodermatitis in breast cancer patients. The results demonstrated a significant delay in radiodermatitis development and progression when treated with silymarin [8]. Unlike skin formulations, oral gels should possess good mucoadhesion, without getting easily washed off by saliva, high water content and low surface friction for better retention in the injured oral mucosa [9]. Although currently, research is ongoing on the utility of topical skin formulation of silymarin, there is no evidence of oral mucoadhesive gel formulation for therapeutic use in various oral diseases. The reason could be attributed to poor water solubility and low bioavailability [2]. However, with the introduction of complexing with phosphatidylcholine, which has better absorption, and new silibinin glyco-conjugates (gluco, manno, galacto, and lacto-conjugates), which both have a high solubility in the water, this may be overcome [10].

Silymarin as an oral topical formulation can aid in the treatment of many oral diseases, taking advantage of its antioxidant, anti-inflammatory, antifibrotic and anticancer activity. To the best of our knowledge, since there is no oral topical formulation available in the literature, the current study aims to design, formulate and evaluate the preclinical release studies of silymarin-based mucoadhesive oral topical gel.

### **2. Results**

### *2.1. Characterization of Silymarin Gel Formulation*

The following findings were obtained from the characterization studies, which gave an overview of the physical properties of the chosen silymarin gel formulation (F10) (Figure 1 and Table 1). The various gel formulations tested in various compositions have been listed in Table S1. *Gels* **2023**, *9*, x FOR PEER REVIEW 3 of 15

**Figure 1.** Silymarin mucoadhesive gel**. Figure 1.** Silymarin mucoadhesive gel.

**S.No Parameters Observation** 

Linear regression analysis It obeys Beers-Lamberts law FTIR spectroscopy No interaction was observed

Viscosity 3700 ± 0.98 to 7400 ± 0.32 cps Homogeneity No visible particles are seen

Taste: Bitter

Stability No change in physical properties was observed

ter 3 h

Release kinetics Zero order, first order, Higuchi kinetic *In vitro* antioxidant Presence of antioxidant activity is confirmed

Odor: Characteristic

Color: Pale yellow color Odor: Mint odor Taste: Astringent taste

*Ex vivo* diffusion study In 3 h, it was found to be 3.03% and coefficient range was 0.9701

for silymarin when analyzed at 287 nm with an R2 = 0.9962.

Percentage cumulative drug release was found to be 2.6% and 3.25% for open-ended cylinder and diffusion cell, respectively, af-

The scanning range ultraviolet spectrophotometric analysis was performed in distilled water, and 287 nm was used as the experimental value of maximum. The absorbance value of standard concentrations of 1–5 μg/mL was plotted, and linearity was observed

**Table 1.** Various parameters evaluated in the chosen silymarin gel formulation (F10)**.** 


**Table 1.** Various parameters evaluated in the chosen silymarin gel formulation (F10).

The scanning range ultraviolet spectrophotometric analysis was performed in distilled water, and 287 nm was used as the experimental value of maximum. The absorbance value of standard concentrations of 1–5 µg/mL was plotted, and linearity was observed for silymarin when analyzed at 287 nm with an R<sup>2</sup> = 0.9962. *Gels* **2023**, *9*, x FOR PEER REVIEW 4 of 15

### *2.2. FTIR Spectral Analysis 2.2. FTIR Spectral Analysis*

ents**.** 

2.3.1. pH

(Table 2).

4.9 ± 0.04

15.98 ± 0.03

3490 ± 0.07

0.5 ± 0.21 (Silymarin: 0.5 g)

6.5 ± 0.12

**Evaluation Parameters** 

**pH** 4.2 ±

**Spreadability** 10.73 ±

**Viscosity (cpm)** 3700 ±

**Stability pH** 6.5 ±

**Theoretical drug content (mg /g)** 

0.02

0.01

0.98

0.2 ± 0.01 (Silymarin: 0.5 g)

0.03

found to be 6.4

2.3.2. Homogeneity

5.0 ± 0.01

18.01 ± 0.02

5000 ± 0.56

0.87 ± 0.78 (Silymarin :0.5 g)

6.5 ± 0.15

*2.3. Evaluation Parameters* 

The FTIR spectra of the pure drug showed prominent peaks at 3566.7 cm−<sup>1</sup> due to O-H stretch, 2925.48 cm−<sup>1</sup> due to C-H stretch, 2.360.44 cm−<sup>1</sup> due to absorption of carbon dioxide, 1716.34 cm−<sup>1</sup> due to C=O stretch cyclic ketone, 1684.52 cm−<sup>1</sup> due to C=C stretch aromatic, 1508.06 cm−<sup>1</sup> due to C-H bending, 1281.47 cm−<sup>1</sup> due to C-OH stretch, and 1166.72 cm−<sup>1</sup> due to C-O stretch. From the spectra (Figure 2), it was observed that there was no significant change in the original peak of the drug and the polymer when compared with the spectra of the physical mixture of the formulated gel. This indicates that there was no interaction between drug, polymer and other excipients. The FTIR spectra of the pure drug showed prominent peaks at 3566.7 cm−1 due to O-H stretch, 2925.48 cm−1 due to C-H stretch, 2.360.44 cm−1 due to absorption of carbon dioxide, 1716.34 cm−1 due to C=O stretch cyclic ketone, 1684.52 cm−1 due to C=C stretch aromatic, 1508.06 cm−1 due to C-H bending, 1281.47 cm−1 due to C-OH stretch, and 1166.72 cm−1 due to C-O stretch. From the spectra (Figure 2), it was observed that there was no significant change in the original peak of the drug and the polymer when compared with the spectra of the physical mixture of the formulated gel. This indicates that there was no interaction between drug, polymer and other excipients.

**Figure 2.** FTIR spectra of the formulation ingredients such as silymarin, carbopol and other excipi-0.04 7.0 ± 0.06 **Figure 2.** FTIR spectra of the formulation ingredients such as silymarin, carbopol and other excipients.

There were no lumps or grittiness in any of the gel formulations that were made.

6.0 ± 0.01

20.56 ± 0.03

6302 ± 0.33

12.1 ± 0.80 (Silymarin :1 g)

6.8 ± 0.31

6.2 ± 0.04

21.64 ± 0.02

6589 ± 0.54

17.20 ± 0.37 (Silymarin :1 g)

6.9 ± 0.01

6.3 ±

22.05 ± 0.04

7000 ± 0.94

29.83 ± 0.25 (Silymarin :1 g)

7.0 ±

0.03 6.4 ± 0.01

23.75 ± 0.03

7400 ± 0.32

32.77 ± 0.20 (Silymarin: 1 g)

**Table 2.** Physicochemical evaluation for various gel formulations.

**Homogeneity** good Good good good good good good good good good

5.5 ± 0.03

18.95 ± 0.04

5231 ± 0.02

0.95 ± 0.12 (Silymarin :0.5 g)

6.6 ± 0.04

**F1 F2 F3 F4 F5 F6 F7 F8 F9 F10** 

5.8 ± 0.04

19.89 ± 0.01

6000 ± 0.51

10.72 ± 0.76 (Silymarin :1 g)

6.8 ± 0.40

5.7 ± 0.05

19.73 ± 0.09

5409 ± 0.34

1.9 ± 0.10 (Silymarin :0.5 g)

6.8 ± 0.21
