Determination of Oligomeric Proanthocyanidins (OPCs) from Tamarindus indica Seed Coat (of Indian origin) using LC/MS/MS

Significance of the Topic

Oligomeric proanthocyanidins (OPCs) from Tamarindus indica seed coats represent a valuable source of natural antioxidants and bioactive compounds. Seed coats are commonly discarded as agro-industrial waste, yet they contain catechin oligomers with potent free-radical scavenging activity, vascular protective effects and anti-inflammatory properties. Developing sensitive analytical methods to identify and quantify these OPCs supports waste valorization and paves the way for new applications in nutraceuticals and antimicrobial formulations.

Objectives and Study Overview

The primary aim of this study was to establish and validate an LC/MS/MS method for the extraction, separation and structural confirmation of OPC dimers, trimers and tetramers from Indian tamarind seed coats. Secondary objectives included evaluation of antimicrobial activity of both crude aqueous extracts and purified OPC fractions against representative Gram-positive and Gram-negative bacteria.

Methodology

Seed coats were separated from kernels by roasting and manual peeling, then finely ground. A 1:10 (w/v) aqueous extraction at 100 °C for 1 hour yielded a proanthocyanidin-rich solution, which was filtered and subjected to C18 solid phase extraction. Sequential elution with diethyl ether, methanol and ethyl acetate selectively recovered catechins and OPCs. The ethyl acetate fraction was dried under nitrogen and reconstituted for analysis.

Used Instrumentation

• Shimadzu LCMS-8030 triple quadrupole mass spectrometer with ESI(–) interface
• Shim-pack XR ODS II column (100 mm × 3 mm, 2.2 µm)
• UV detector set at 280 nm
• C18 SPE cartridges (100 mg, 3 mL)

Analytical Conditions

• Mobile phase A: water with 1 % acetonitrile and 0.1 % formic acid; B: acetonitrile
• Gradient: 3→9 % B (0–5 min), 9→16 % B (5–15 min), hold 16 % (15–25 min), 16→3 % B (25–26 min)
• Flow rate: 0.4 mL/min; column oven: 40 °C; injection: 20 µL
• MS: desolvation line 250 °C; heat block 400 °C; nebulizing gas 3 L/min; drying gas 15 L/min

Main Results and Discussion

UV chromatograms at 280 nm revealed distinct elution peaks corresponding to OPC oligomers. In negative-ion single ion monitoring, m/z 577, 865 and 1153 signals confirmed the presence of dimer, trimer and tetramer units, respectively. MS/MS fragmentation patterns were consistent with B-type procyanidin linkages. Antimicrobial sensitivity tests using agar ditch and agar cup methods showed that both crude aqueous extract and OPC fraction inhibited growth of Staphylococcus aureus ATCC 6538 P and Bacillus subtilis, whereas Escherichia coli and Salmonella typhi remained unaffected. Notably, the crude extract exhibited larger inhibition zones than the purified OPC fraction, indicating synergistic effects of other seed coat constituents.

Benefits and Practical Applications

• Valorization of tamarind seed coat waste into high-value phytochemicals
• A robust LC/MS/MS protocol for qualitative confirmation of OPCs without the need for pure standards
• Potential development of natural antimicrobial agents targeting Gram-positive pathogens
• Foundations for functional nutraceutical and cosmetic ingredients leveraging antioxidant and vascular protective properties

Future Trends and Potential Applications

• Scale-up of extraction and purification processes for industrial production of OPC concentrates
• Quantitative method validation and calibration strategies for routine quality control
• Exploration of synergy between OPCs and other phytochemicals for broader antimicrobial spectra
• Integration of OPCs into drug delivery systems, wound-healing formulations and dietary supplements
• Application of advanced MS techniques (e.g., high-resolution MS, ion mobility) to resolve structural isomers

Conclusion

The developed LC/MS/MS method successfully identified dimeric, trimeric and tetrameric OPCs in Tamarindus indica seed coat extracts, confirming their structures via characteristic fragmentation pathways. Antimicrobial testing demonstrated selective activity against Gram-positive bacteria, with crude extracts outperforming purified fractions. These findings highlight the potential of tamarind seed coats as a sustainable source of bioactive OPCs and support further research into their practical applications.

Reference

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