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INTRODUCTION
Arthritic pain is currently one of the major burning issues in our society, with a total of 18 million people worldwide suffering from Rheumatoid Arthritis (RA) in 2019. This would translate into a total of almost seven million patients in India if projected to the entire population (Malaviya et al., 1993). The prevalence of RA in adults has been reported to vary from 0.5 to 3.8% in women and from 0.15 to 1.37% in men, with peak incidence in the fourth decade of life (Lawrence, 1977). It can affect any synovial joint in the body, most commonly starting in the small joints of the hands and feet, with the potential to impact every aspect of daily living. High levels of inflammation are associated with fatigue and impairment of participation in occupational, recreational, and societal roles (GBD, 2021 Rheumatoid Arthritis Collaborators, 2023). The pathogenesis and clinical characteristics of RA do not match any specific Ayurvedic disease. However, Vataraktam, rheumatism due to Vata and Rakta (NAMC Code ED-8), covers various musculoskeletal and rheumatological disorders, along with conditions affecting the skin and vasculature. Vataraktam (polyarthritis caused by Vata and Rakta) is classified by the WHO Standard Terminologies (ITA-5.30) as a disorder that aligns with RA based on its symptoms. (Acharya, 2006) Guduchi Taila is one of the formulations mentioned in various Ayurveda texts, such as Sushruta Samhita, Ashtanga Hridaya, Chakradutta and Vangasena in the management of Vataraktam (Sharma et al., 2025). This Guduchi Taila is processed for one hundred cycles through a process called Aavartana, to enhance its bioavailability, reduce the required dosage, and provide Rasayana (rejuvenative and therapeutic) effects, to finally get the formulation Shatapaka Guduchi Taila (100 Times Processed Guduchi Taila) (hereafter SGT).
Gas Chromatography-Mass Spectrometry (GC-MS) is a technique ideal for metabolomic profiling of vaporised single or a blend of various plant samples (Kanthal et al., 2014). A wide range of phytochemical compounds can be sought with a single run of the sample based on its nature. The obtained metabolites are identified through mass spectrometry based on their mass-to-charge ratio (Venkataramanam et al., 2023). Despite the prominent mention of Shatapaka Guduchi Taila across Ayurvedic classical treatises and its recognised therapeutic importance, a systematic GC–MS-based phytochemical profiling of the formulation has not yet been documented. This study aims to quantitatively analyse the phytochemical compounds present in SGT through the GC-MS analysis. Despite its traditional relevance in the management of Vataraktam, the broader therapeutic scope of SGT and its mechanism of action in classically enlisted disease have not been comprehensively investigated. The current study represents a preliminary attempt to expand this understanding.
MATERIALS AND METHODS
Study Conduction
The Shatapaka Guduchi Taila was prepared at The Indian Medical Practitioner’s Co-operative Pharmacy Stores Ltd. (IMPCOPS) Chennai-41, a GMP-certified Ayurveda Pharmacy, as per Ayurveda Pharmacopeia standards. Table 1 shows the ingredients with proportions used for the preparation of SGT. The authentication of drugs and physicochemical analysis of SGT were done at IMPCOPS Ltd., Chennai. The prepared sample was analysed for organoleptic parameters, including colour, odour, taste, and consistency, as well as physico-chemical analysis detailed in Table 2. The GC-MS analysis was done at the Centre for Analytical Instrumentation-Kerala (CAI-K), Kerala Forest Research Institute (KFRI) at Peechi, Thrissur District, Kerala.
| Sl. No. | Ingredient | Drug | Latin Name | Part Used | Proportion |
|---|---|---|---|---|---|
| 1 | Kalka Dravya (Paste Drug) | Guduchi | Tinospora cordifolia [Willd.] Miers | Stem | ¼ Parts |
| 2 | Sneha Dravya (Oil base) | Tila Taila | Sesamum indicum | Oil | 1 Part |
| 3 | Drava Dravya (Kashaya-Decoction) | Guduchi Kashaya | Tinospora cordifolia [Willd.] Miers | Stem | 4 Part |
| 4 | Avapa Drava Dravya (Added Drug) | Goksheera | Cow milk | Milk | 1 part |
| Parameters | Characters |
|---|---|
| Description | Pale yellow white colour, oily, unctuous, viscous liquid; ghee-like odour; taste- bitter taste |
| Colour under UV Rays | Whitish blue |
| Colour with solvent under UV (Petroleum ether) | Whitish blue |
| Loss on drying at 105°C | 0.2693% |
| Refractive index | 1.4609 @ 24.9°C |
| Specific gravity | 0.9152 |
| Acid value | 1.4655 mg/KOH |
Instrument
GC-MS analysis of SGT was carried out using a Shimadzu GCMS-QP2020 system, which is extensively employed for accurate qualitative and quantitative analysis due to its high sensitivity and analytical reliability.
Sample Preparation
Hexane Extraction-1 µL of hexane was used to extract 1 g of the taila sample. A clear solution was obtained after the removal of solid residue by filtering the extract.
Procedure
A 1 µL aliquot of the SGT sample was dissolved appropriately in HPLC-grade hexane, and 1 µL of this solution was injected into the system in split mode (split ratio 20:1). Helium was employed as the carrier gas at a constant flow rate of 1.0 mL/min. The injector temperature was maintained at 280°C. The oven temperature program was set as follows: initial temperature 70°C, ramped at 8°C/min to 260°C with a 2-min hold, followed by a second ramp of 4°C/min to 280°C with a final hold of 5 min. The ion source temperature was maintained at 220°C, and the interface temperature at 280°C. A solvent cut time of 3.10 min was applied. Mass spectra were recorded in scan mode over an m/z range of 50–500 with a scan speed of 1666 scans/s. Identification of analytes was carried out by comparing mass spectral fragmentation patterns with those in the NIST 2020 library, and compounds were accepted based on high Similarity Index (SI) scores and consistency of characteristic ions.
Ethical Statement
Ethical approval was not required for this study as it involved only the phytochemical analysis of a herbal formulation and did not involve human or animal subjects.
Statistical Analysis
There is no specific subsection for statistical analysis. The study relies on quantitative analysis (area percentage) and Similarity Indices (SI ≥90%) rather than hypothesis testing statistics.
RESULTS
The Total Ion Chromatograph (Figure 1) shows 12 major peaks between 6.03 and 35.24 min. The most abundant compound is Cholesterol (49.06% area) at RT 34.292 min. Second major compound: 2,6-Bis(3,4-methylenedioxyphenyl)-3,7-dioxabicyclo (3.3.0) octane (32.69%) at RT 35.246 min. The name of the compounds detected, their retention time, area, height, area/height ratio and structure has been detailed in Table 3.
| Sl. No. | Name of the compound | Retention time | Area | Area% | Height | Height% | A/H | Structure | Chemical Formula | Compound Class | Activity of Compound |
|---|---|---|---|---|---|---|---|---|---|---|---|
| 1 | Maltol | 6.034 | 104767 | 1.29 | 33543 | 2.79 | 3.12 | C₆H₆O₃ | Phenol Derivative | Antioxidant (Song et al., 2015), Anticancer, (Han et al., 2023), Anti-Inflammatory (Li et al., 2024), Hepatoprotective (Liu et al., 2018), Cardioprotective (Xing et al., 2022). | |
| 2 | 2-Decenal, (E) | 8.523 | 50282 | 0.62 | 19158 | 1.59 | 2.62 | C₁₀H₁₈O | Unsaturated Aldehyde | Antileishmanial (Donega et al., 2014), Anticancer (Vijayakumar et al., 2025). | |
| 3 | 2H-Pyran-2-one, tetrahydro-6-nonyl- | 18.301 | 24109 | 0.3 | 13062 | 1.09 | 1.85 | C₁₄H₂₆O₂ | Tetrahydropyranone | Antibacterial, Antiviral, and Anti-Inflammatory (Nazari et al., 2019), Anti-Alzheimer's (Almalki et al., 2023). | |
| 4 | 2H-Pyran-2-one, tetrahydro-6-tridecyl | 20.866 | 41957 | 0.52 | 16268 | 1.35 | 2.58 | C₁₈H₃₄O₂ | Tetrahydropyranone | Anticancer And Antioxidant (Mashrai et al., 2013). | |
| 5 | 9-Octadecenoic acid (Z)-, oxiranylmethyl ester | 24.511 | 69919 | 0.86 | 29088 | 2.42 | 2.4 | C₂₁H₃₈O₃ | Fatty Acid Ester | Antioxidant, Anti-Inflammatory, Antimicrobial & Cytotoxic (Sharaf et al., 2021). | |
| 6 | Squalene | 29.149 | 124916 | 1.54 | 40722 | 3.39 | 3.07 | C₃₀H₅₀ | Triterpene Hydrocarbon | Hypocholesterolemic (Hien et al., 2017), Anti-Inflammatory and Anti-Cancer (Abuobeid et al., 2022). | |
| 7 | (R)-6-Methoxy-2,8-dimethyl-2-((4R,8R)-4,8,12-trimethyltridecyl) chroman | 32.922 | 103652 | 1.28 | 27459 | 2.29 | 3.77 | C₂₉H₅₀O₂ | Tocopherol/ Chroman Derivative | Anti-Cancerous (Das et al., 2016; Saavedra et al., 2020; Birringer et al., 2003), Antiepileptic (Rawat et al., 2016), Antioxidant, (Lakkadi et al., 2024), Anti-Inflammatory (Jiang et al., 2014; Reiter et al., 2007). | |
| 8 | Beta-Sitosterol acetate | 33.704 | 158159 | 1.95 | 32422 | 2.7 | 4.88 | C₃₁H₅₂O₂ | Sitosterol | Anti-Inflammatory, Antioxidant (Hidayathulla et al., 2018), Anti-Gastro-ulcer (Xiao et al., 1992), Analgesic (Villasenor et al., 2002). | |
| 9 | Cholesterol | 34.292 | 3982353 | 49.06 | 558685 | 46.51 | 7.13 | C₂₇H₄₆O | Cholestanoids | Immune Regulation, Inflammatory (Hien et al., 2017). | |
| 10 | Laurin, 2-capri-1,3-di- | 34.292 | 425863 | 5.25 | 74662 | 6.22 | 5.7 | C₃₇H₇₀O₉ | Glycerol Ester | Antimicrobial, (Matsue et al., 2019), Antioxidant (Ameena et al., 2024). | |
| 11 | 3-(Octanoyloxy) propane-1,2-diyl bis(decanoate) | 35.065 | 376975 | 4.64 | 89862 | 7.48 | 4.2 | C₃₁H₅₈O₉ | Triglycerols | Anti-Inflammatory, Anti-Oxidant (Ratheesh et al., 2022). | |
| 12 | 2,6-Bis (3,4- methylenedioxyphenyl)-3,7-dioxabicyclo (3.3.0) octane | 35.246 | 2653550 | 32.69 | 266198 | 22.16 | 9.97 | C₂₀H₁₈O₆ | Phenylpropanoid Dimers | Antioxidant (Nakai et al., 2003). |
DISCUSSION
Shatapaka Guduchi Taila is prepared based on the Chakradutta Vataraktam Chikitsa reference, utilising Guduchi (Tinospora cordifolia [Willd.] Miers), Tila Taila (Sesamum indicum L.), and Goksheera (Cow Milk). Guduchi Taila is an oleaginous compound formulation that comprises Guduchi (Kalka Dravya), Tila Taila (Sneha Dravya), and Goksheera (Drava Dravya). It is prepared as per the principles of Sneha Kalpana as outlined in the Sharangdhara Samhita. In Ayurveda, there exists a methodology known as Aavartana, which involves the repeated processing of substances. The Guduchi Taila is processed for one hundred cycles (Aavartana) to enhance its bioavailability, reduce the required dosage, and provide Rasayana (rejuvenative and therapeutic) effects, to get Shatapaka Guduchi Taila (Sharma et al., 2024). Guduchi is regarded as the best drug for the management of Vataraktam (ED-8 Rheumatoid Arthritis) (Sharma et al., 2024). Multiple formulations have been described in Ayurveda texts that contain Guduchi as the main ingredient or as a constituent. There are 98 references available for the indication of Guduchi in Vataraktam, where the maximum used dosage form is Kashaya/Kwatha (decoction) with 41 formulations (Sharma et al., 2024; Shedbale et al., 2025). Properties of Tila Taila help to combat the vitiated Vata and Rakta. (Bhavaprakasha Nighantu, n.d.; Madanapala Nighantu n.d.) The ushna (hot) veerya (potency) pacifies the Vata, and madhura vipaka (biotransformation) works on Rakta vitiation. It also possesses properties of balya (strengthening), chakshushya (promotes eyesight), Deepana (promotes digestion), keshya (strengthens hair), etc., (Sharma et al., 2024). Goksheera (cow milk) is commonly utilised as a dietary source in all age groups. Ayurveda advises Goksheera be taken daily and regards it as the Rasayana (rejuvenating) based upon its properties (Sharma et al., 2025). It pacifies the Vata and Pitta Dosha, Vata and Rakta, enhances the complexion, and is an immuno-regulator (Sharma et al., 2024). This mode of action, which is based on rasa, guna, veerya, vipaka, and karma, supports the efficacy of SGT in treating Vataraktam on the lines of Ayurveda fundamentals. Moreover, this study aims to provide evidence based on the mode of action of phytochemical compounds, thereby contributing to the validation of traditional knowledge in the context of ED-8 Rheumatoid Arthritis and other related disorders.
The GC-MS analysis of Shatapaka Guduchi Taila reveals 12 total compounds, as listed in Table 3. This analysis of the sample revealed a complex phytochemical and lipid-based profile comprising aldehydes, lactones, fatty acid esters, terpenoids, sterols, and polyphenolic derivatives. Compound identification was accomplished through comparison with the NIST 20 mass spectral library, yielding high similarity indices (SI ≥ 90), thereby confirming the reliability of identification. The predominance of bioactive compounds with reported antioxidant, anti-inflammatory, immunomodulatory and chemo-preventive properties suggest potential pharmacological relevance of the analysed sample. The common activity of the compounds is reported in Table 4.
| Sl. No. | Activity | Number | Name of the Compound |
|---|---|---|---|
| 1 | Anti-Oxidant | 6 | Maltol; 9-Octadecenoic acid (Z)-, oxiranylmethyl ester; (R)-6-Methoxy-2,8-dimethyl-2-((4R,8R)-4,8,12-trimethyltridecyl) chroman; Beta-Sitosterol acetate; 3-(Octanoyloxy) propane-1,2-diyl bis(decanoate); 2,6-Bis(3,4-methylenedioxyphenyl)-3,7-dioxabicyclo (3.3.0) octane |
| 2 | Anti-Cancerous | 4 | 2-Decenal, (E); Maltol; 2H-Pyran-2-one, tetrahydro-6-tridecyl; Squalene |
| 3 | Anti-Inflammatory | 7 | Maltol; 2H-Pyran-2-one, tetrahydro-6-nonyl-; 9-Octadecenoic acid (Z)-; oxiranylmethyl ester; Squalene; (R)-6-Methoxy-2,8-dimethyl-2-((4R,8R)-4,8,12-trimethyltridecyl) chroman; Beta-Sitosterol acetate; 3-(Octanoyloxy) propane-1,2-diyl bis(decanoate) |
| 4 | Anti-Bacterial | 4 | Laurin, 2-capri-1,3-di-; 9-Octadecenoic acid (Z)-; oxiranylmethyl ester |
In this GC-MS analysis, Cholesterol, 2,6-Bis(3,4-methylenedioxyphenyl)-3,7-dioxabicyclo (3.3.0) octane, Laurin, 2-capri-1,3-di-, and Beta-Sitosterol acetate, has the highest area and height ratio, which is linked to the highest concentration of the phytochemical in the compound in SGT.
Cholesterol is essential for cellular integrity, immune regulation, and inflammatory balance. It maintains membrane stability and signalling and serves as a precursor for steroid hormones, bile acids, and vitamin D, supporting physiological homeostasis (Cortes et al., 2014). Balanced cholesterol levels regulate immune cell activity, whereas dysregulated metabolism promotes macrophage activation and chronic inflammation (Tall et al., 2015; Aguilar-Ballester et al., 2020). In rheumatoid arthritis and related inflammatory joint disorders, cholesterol imbalance is associated with disease progression and inflammatory burden (Lei et al., 2023; Robinson et al., 2022). From an Ayurvedic perspective, these functions are parallel to sneha and meda dhatu metabolism, which supports Vata alleviation and joint lubrication (sandhi snehana) (Agnivesha, 1983). Thus, physiological cholesterol balance can help in inflammatory modulation in Vataraktam-like conditions, while excess or oxidised cholesterol may contribute to Rakta vitiation and disease aggravation (Susruta, 2001; Agnivesha, 1983).
2,6-Bis(3,4-methylenedioxyphenyl)-3,7-dioxabicyclo (3.3.0) octane
A furofuran lignan, has demonstrated antioxidant activity due to its catechol-like structure. Related sesamin metabolites with the same 3,7-dioxabicyclo (3.3.0) octane core effectively scavenge reactive oxygen species, highlighting their potential to modulate oxidative stress (Nakai et al., 2003). Since oxidative stress contributes to rheumatoid arthritis pathogenesis by promoting inflammation and joint damage, antioxidants like this compound may help reduce inflammatory cascades and tissue injury in RA (Robinson et al., 2022).
Laurin, 2-capri-1,3-di- (Monolaurin (glycerol monolaurate)), a derivative of lauric acid, has demonstrated broad-spectrum antimicrobial activity against Gram-positive and Gram-negative bacteria isolated from skin infections, with strong sensitivity and low resistance rates in vitro (Carpo et al., 2007). Also showing inhibition of antibiotic-resistant Staphylococcus aureus in atopic dermatitis without cytotoxicity, suggesting potential value in treating resistant skin disorders (Laowansiri et al., 2025). Additionally, it exhibits antibiofilm activity, which is critical for managing chronic or wound-associated bacterial colonisation (Hassan et al., 2024). Although no direct studies have examined monolaurin in rheumatoid arthritis, the pathophysiology of RA involves significant oxidative stress and immune dysregulation, and antioxidant interventions have shown moderate benefits in reducing oxidative damage and inflammation in RA contexts (Djordjevic et al., 2023). These findings support the broader concept that compounds combining antimicrobial and antioxidant activities may have relevance in inflammatory and immune-mediated disorders.
Beta-Sitosterol acetate or β-Sitosterol
A common plant phytosterol, has widely reported its antiinflammatory and antioxidant properties. In cellular and animal models, it has shown suppression of key inflammatory mediators such as TNFα, IL1β, and IL6, inhibit inflammasome activation and NFκB signalling, and reduces reactive oxygen species, demonstrating its capacity to downregulate inflammatory responses and oxidative stress in vitro and in vivo. It reduced oxidative stress and neutrophil recruitment in a zebrafish inflammation model by increasing antioxidant enzymes like sod and gpx4b, while decreasing pro-inflammatory genes such as il-8 and myd88, highlighting its strong antioxidant and anti-inflammatory in inflammatory disorders (Zhang et al., 2023). Additionally, in RA, βsitosterol has exhibited antiarthritic actions by inhibiting synovial angiogenesis, reducing joint swelling and cartilage damage, suppressing VEGF signalling, and improving redox balance in collageninduced arthritis models, suggesting therapeutic potential for RA. These findings support the idea that β-sitosterol and its derivatives may mitigate inflammation and oxidative stress mechanisms involved in RA pathogenesis (Qian et al., 2022).
Antioxidant Potentials of Identified Compounds
Oxidative stress plays a crucial role in the pathogenesis of metabolic and chronic inflammatory disorders like RA and cancer. Several compounds identified in GC–MS analysis of SGT are well documented for their antioxidant properties. Maltol, a pyranone derivative detected in the early elution phase, is known for its potent free radical scavenging ability and inhibition of lipid peroxidation (Halliwell et al., 1994). Squalene, a triterpenoid hydrocarbon identified with high confidence, functions as a chain-breaking antioxidant that protects cellular membranes from oxidative damage (Hien et al., 2017; Abuobeid et al., 2022). Tocopherol-related chroman derivatives identified in the sample further enhance antioxidant defence by neutralising Reactive Oxygen Species (ROS) and preventing oxidative degradation of polyunsaturated fatty acids (Lakkadi et al., 2024). Methylenedioxyphenyl-containing compounds, which formed a significant proportion of the chromatographic area, are reported to exert strong antioxidant effects through modulation of redox-sensitive pathways and metal chelation. Collectively, the abundance of these antioxidant constituents suggests that the sample possesses substantial oxidative stress-modulating potential, which is particularly relevant in inflammation-driven disorders (Halliwell et al., 2006).
Anti-Inflammatory Potentials of Identified Compounds
Inflammation is a central pathological feature underlying rheumatoid arthritis, and other metabolic diseases. Phytosterols such as β-sitosterol acetate identified in the present study are widely reported to suppress inflammatory mediators, including Cyclooxygenase-2 (COX-2), TNF-α, and interleukins (Loizou et al., 2010; Vilahur et al., 2019; Marahatha et al., 2021). Squalene has also been shown to attenuate inflammatory signalling by modulating NF-κB activation and reducing oxidative inflammation (Ibrahim et al., 2021). Additionally, lactone derivatives such as tetrahydro-2H-pyran-2-one analogues contribute to anti-inflammatory activity by inhibiting nitric oxide synthesis and prostaglandin release. The cumulative presence of these compounds suggests that the sample may exert multi-targeted anti-inflammatory effects, supporting its relevance in chronic inflammatory conditions (Gan et al., 2019).
Anticancer and Chemopreventive Potentials of Identified Compounds
Several compounds identified in the GC-MS profile of SGT, like Maltol, 2-Decenal, (E)-, Squalene, 2H-Pyran-2-one, and tetrahydro-6-tridecyl have been associated with anticancer or chemopreventive activities. Squalene has been extensively studied for its ability to inhibit tumour progression, enhance immune surveillance, and reduce oxidative DNA damage (Abuobeid et al., 2022). Methylenedioxyphenyl derivatives are structurally linked to apoptosis induction and inhibition of carcinogen-activating enzymes. Tocopherol analogues further contribute by protecting cellular DNA from oxidative mutations (Woyengo et al., 2009). Although direct anticancer activity cannot be concluded from chemical profiling alone, the presence of these compounds provides a scientific basis for further in vitro and in vivo investigations aimed at evaluating the chemopreventive potential of the sample.
In summary, the GC-MS analysis of compounds of SGT, collectively confers antioxidant, anti-inflammatory, antimicrobial, and potential chemopreventive benefits, aligning with Ayurvedic principles of Vataraktam modulation while paralleling ED-8 RA pathogenesis management through ROS scavenging, cytokine suppression, and membrane stabilisation. This phytochemical synergy positions SGT as a promising candidate for further validation in inflammatory disorders, bridging traditional formulation wisdom with evidence-based therapeutic potentials.
CONCLUSION
The GC-MS profiling of Shatapaka Guduchi Taila (SGT) identified 12 bioactive compounds, predominantly cholesterol (49.06%), 2,6-Bis(3,4-methylenedioxyphenyl)-3,7-dioxabicyclo (3.3.0) octane (32.69%), and β-sitosterol acetate, revealing a rich lipid matrix with antioxidant, anti-inflammatory, anti-cancer and antimicrobial potentials. These phytoconstituents align with Ayurvedic principles of Vataraktam management through sneha guna and dhatu nourishment, paralleling RA pathogenesis management via ROS scavenging, cytokine suppression, immuno-modulation and membrane stabilisation. SGT thus emerges as a promising Rasayana (rejuvenative and therapeutic) candidate bridging classical wisdom with evidence-based rheumatological therapeutics, warranting advanced in vitro/in vivo validation.