The Study of Optimal Solvent Conditions for the Extraction of Phyllanthus Emblica L. Using Ultrasonic-Assisted Extraction

Abstract

Introduction and Objective: Phyllanthus emblica L., commonly known as Indian gooseberry or "Makham Pom" in Thailand, is a medicinal plant widely recognized for its therapeutic potential. This fruit has played a significant role in both Thai traditional medicine and Ayurvedic practices for centuries. Traditionally, it has been incorporated into remedies for various respiratory conditions, especially in formulations aimed at alleviating cough, soothing irritated throats, reducing phlegm production, and enhancing immune system function. The efficacy of P. emblica is attributed to its high content of bioactive constituents, including but not limited to kaempferol, gallic acid, tannins, and ascorbic acid (vitamin C). These compounds have demonstrated a wide range of pharmacological activities such as antioxidant, anti-inflammatory, anticancer, antidiabetic, and antiviral properties. Despite its well-documented medicinal properties, the extraction techniques conventionally employed to isolate these compounds such as decoction, boiling, and maceration with water or ethanol present several drawbacks. These traditional approaches are time consuming and often involve high temperatures or prolonged exposure to solvents, potentially causing degradation or transformation of thermolabile and pH sensitive phytochemicals. Such limitations underscore the necessity for developing and implementing more efficient, selective, and sustainable extraction technologies that preserve the integrity and biological activity of the target compounds.

     Ultrasonic-assisted extraction (UAE) has gained considerable attention in recent years as an effective and eco-friendly alternative to conventional extraction methods. This technique utilizes high-frequency ultrasonic waves to induce cavitation, thereby enhancing mass transfer, cell disruption, and solvent penetration. UAE not only shortens extraction time and improves yield but also reduces the need for harsh solvents and elevated temperatures. Consequently, this technique is highly suitable for the extraction of bioactive molecules from natural sources, particularly when targeting compounds that are sensitive to heat and oxidation. The present study aimed to evaluate the impact of different solvent systems on the efficiency of UAE in extracting key bioactive compounds from P. emblica. Specifically, the objective was to determine which solvent yields the highest levels of antioxidant activity and phytochemical content, particularly gallic acid, tannins, and vitamin C. The outcomes of this research are expected to contribute valuable insights into the optimization of extraction protocols for pharmaceutical, nutraceutical, and cosmeceutical applications.

     Methods: Fresh fruits of P. emblica used in this study were sourced from cultivated orchards in Kanchanaburi Province, Thailand., thoroughly washed to remove impurities, and subsequently dried in a hot air oven at 50 °C for 72 hours. This drying temperature was chosen to effectively reduce the moisture content while preserving the structural integrity of heat-sensitive phytochemicals. The dried fruits were ground into fine powder using a mechanical grinder. The powdered material was then subjected to ultrasonic-assisted extraction using four different solvent systems: distilled water, 50% ethanol, 70% ethanol, and 90% ethanol. Each extraction was performed using an ultrasonic processor operating at a frequency of 20 kHz for 60 minutes at room temperature. Post-extraction, the liquid extracts were filtered, and the solvents were removed via freeze-drying (lyophilization), yielding powdered extracts for subsequent analysis. The qualitative identification of major phytochemicals, namely gallic acid and ascorbic acid, was conducted using High-Performance Thin Layer Chromatography (HPTLC). Chromatograms were developed and visualized under white and UV (366 nm) light, and retention factor (Rf) values were compared against standard reference compounds. Quantitative assessment of gallic acid, total tannin content, and ascorbic acid was performed using High-Performance Liquid Chromatography (HPLC), following validated analytical protocols. Antioxidant capacity of the extracts was determined using the DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging assay.

     Results: HPTLC analysis successfully confirmed the presence of gallic acid and ascorbic acid in all extracts, regardless of the solvent system used. Gallic acid was observed as a grayish-brown band with an Rf value of 0.40 under white light and exhibited a bright blue fluorescence under UV light, matching the standard. Ascorbic acid appeared as a light brown band at an Rf value of 0.16 under white light and a dark brown hue under UV light, consistent with the standard reference. Quantitative results revealed a significant variation in extraction yield and phytochemical content depending on the solvent employed. Distilled water yielded the highest extract mass, with a yield of 26.81 ± 0.30%, and also contained the highest concentration of ascorbic acid, measured at 935.57 ± 0.96 mg/100g. These values were significantly greater (p < 0.05) than those observed in ethanol based extracts. Conversely, 90% ethanol was the most effective solvent for extracting gallic acid and tannins. This solvent yielded the highest concentrations of gallic acid 13.16 ± 0.01 g/100 g, total tannins 208.89 ± 0.26 mg TAE/g, and antioxidant activity 122.60 ± 1.35 g eq ascorbic acid/100 g, with statistical significance (p < 0.05).

     Discussion: Ethanol 90% was the most suitable solvent for the ultrasonic-assisted extraction of P. emblica in this study, as it provided the highest content of gallic acid, tannin and antioxidant activity. However, the extraction with distilled water gave the highest extraction yield and vitamin C.

     Conclusion and Recommendation: Ethanol 90% is the ideal solvent for ultrasonic-assisted extraction of P. emblica in this study because it exhibited the high amounts of active compounds and good antioxidant activity. This study can serve as a guideline for manufacturing the P. emblica extracted to application in Thai Traditional Medicine and health products. Nonetheless, the aqueous extraction of P. emblica remains a viable method if the emphasis is on harnessing vitamin C from the extract.