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Tea was planted in Lahijan by Kashefalsataneh in 1930. The main concern about important commercial plants such as tea is the formation of ice crystals in low temperatures. This can damage the live cells leading to lowering the quality of the plant and eventually its death. Formation of reactive oxygen species (ROS) and oxidative stress is the result of various environmental stresses leading to freezing. Investigating the variations in any of these factors could help to understand the mechanism of freeze resistance in ever-green plants. The aim of the present research was to investigate lipid peroxidation, the presence of antifreeze protein and variations in the activity of some antioxidant enzymes, including superoxide dismutase (SOD), ascorbate peroxidize (APX) and catalyse (CAT) in tea leaves subjected to 20, 0, -2, -5 and -8°C in tea leaves from the north of Iran. The results showed formation of an antifreeze protein with MW of about 20 KD in response to cold stress. It was also found that the activity of SOD, APX and CAT increased in tea leaves due to cold stress. The activity of SOD increased down to -8°C. APX and CAT increased their activity down to -5°C. On the other hand, the lipid per oxidation factor, MDA, was also elevated in response to the cold stress.

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Peroxidase catalyzes different oxidation of substrates using hydrogen peroxide, a reactive oxygen specie (ROS). ROS, at low concentrations, act as messenger to regulate intracellular signaling, whereas, at high concentrations, they can overcome the immune system by creating oxidative stress. Some common beverages such as coffee, tea and soft drinks contain high levels of xanthine alkaloids including theophylline and theobromine. In this study, the effect of theophylline and theobromine on peroxidase activity was kinetically studied by measuring the absorption of 4-aminoantipyrine, oxidized in the presence and absence of theophylline and theobromine at 510 nm for 3 minutes. The results showed that theobromine and theophylline acted as inhibitors with IC₅₀ of 0.50 and 0.55 mM, respectively. K_m and V_max values showed that both compounds are non-competitive inhibitors. The values of K_i were calculated as 0.03 and 0.045 mM for theobromine and theophylline, respectively. Lower values of K_i and IC₅₀ for theobromine compared to theophylline indicates that theobromine has a higher inhibition strength and binding tendency to the enzyme-substrate complex. Hence, it is concluded that theobromine has a stronger inhibitory effect on POD activity.