Abstract:

The interaction between purple sweet potato anthocyanins and trypsin was studied by measuring the catalytic
activity and reaction kinetics through ultraviolet (UV) absorption, fluorescence and infrared (IR) spectroscopy. The
results showed that purple sweet potato anthocyanins had an obvious inhibitory effect on trypsin catalytic activity, and the
inhibition was reversible competitive inhibition with an inhibitory constant (Ki) of 6.16 × 10-4 mmol/L. When the reaction
between purple sweet potato anthocyanins and trypsin with a mole ratio of 140:1 was carried out at 37 ℃ for 15 min,
the inhibitory rate was 38.61%; however, the catalytic activity was not obviously affected by reaction time. Purple sweet
potato anthocyanins treatment led to the quenching of intrinsic fluorescence of trypsin. The quenching of trypsin by purple
sweet potato anthocyanins was probably a static quenching process with a quenching constant (Kq) of 1.73×1012 L/(mol·s),
and a binding constant (KA) of 3.88×104 L/mol. The number of binding sites was 0.86. According to the thermodynamic
parameters, hydrogen bonding and van der Waals force played a dominant role in the interaction between the anthocyanins
and trypsin. The distance between donor and acceptor in anthocyanin-trypsin was calculated as 3.56 nm, based on the
equations from Förster non-radiation energy transfer theory. The IR spectra revealed the conformational change of trypsin
caused by binding, thus leading to a decrease of α-helix and an increase of β-fold.

Key words: purple sweet potato anthocyanins, trypsin, enzymatic properties, ultraviolet (UV) absorption spectroscopy, fluorescence spectroscopy, infrared (IR) spectroscopy