Cytotoxicity and 3T3-L1 cell uptake of lactucaxanthin purified and characterized by LC-MS and NMR from lettuce (Lactuca sativa)

Abstract

Lettuce (Lactuca sativa) and its bioactive constituent lactucaxanthin are gaining importance due to their antidiabetic property. Hence, aim of this study is to establish a simple purification technique for lactucaxanthin and its antioxidant property, cytotoxicity, and cellular uptake in 3T3-L1 cells (pre-adipocytes). Results revealed that lactucaxanthin is higher (mg/100 g dry weight) in green lettuce (3.05 ± 0.12) than Romania (0.42 ± 0.05) and Iceberg (0.36 ± 0.06), respectively. The purity of lactucaxanthin separated by TLC followed by preparatory HPLC is 99 ± 1% with λ_max 439 nm. FTIR, LC-MS (M + H⁺‒H₂O = 551) and NMR analysis revealed the characteristic chemical structure of lactucaxanthin. Cytotoxicity of lactucaxanthin in 3T3-L1 cells treated at 1, 3, 5, 10, 30, 50, 60, 100 μM for 24 hr revealed 86–89% cell viability up to 10 µM concentration. 2,2-Diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS)-mediated radical scavenging (35.65 ± 0.85 µg/ml and 4.92 ± 0.81 µg/ml), total antioxidant 5 µg (equivalent to 23.71 ± 1.88 ascorbic acid) activity and reductive capability 20 µg (26.14 µg quercetin equivalent) of lactucaxanthin proved its antioxidant potential. 3T3-L1 cellular uptake of lactucaxanthin was found time dependent with absorption maxima of 46.35 ng after 24 hr treatment (10 µM). The present results help in effective positioning of lactucaxanthin for food and pharma application to fight against diabetes and dyslipidemia.

Graphical Abstract

Keywords:

• Cytotoxicity
• HPLC
• lactucaxanthin
• lettuce
• NMR
• spectroscopy
• TLC

Acknowledgments

J. Naveen (Award No. F./PDFSS-2015-17-KAR-11600) and R. E. Anitha acknowledge the University Grant Commission and Department of Biotechnology, Govt. of India for awarding Post-Doctoral Fellowship and Junior Research Fellowship.

Disclosure statement

The authors declare that they have no conflict of interest.

Additional information

Funding

Authors also acknowledge University Grant Commission (UGC-PDF, India), DST-SERB for financial support (Award No. EMR/2017/002047; dt. 14.11.2018) and Department of Biotechnology and Nutrition Biology (BT/PR10329/PFN/20/776/2013), Ministry of Science and Technology.