Table 3.

Methods of nattokinase activity determination.

Method	Principle	Feature	Reference
Fibrin plate method	Nattokinase hydrolyzes fibrin on the plate to form a hydrolysis circle	The accuracy is not high, only used for preliminary characterization of enzyme activity	[55]
Fibrinolysis method	A standard curve was established using the time taken for urokinase to dissolve a certain amount of fibrin with different activity units.	Simple with limited precision	[56]
Tetrapeptide substrate method	Hydrolysis of Suc-Ala-Ala-Pro-Phe-pNA by nattokinase to produce chromogenic nitroaniline	Simple and fast, high sensitivity; only suitable for pure enzymes	[57]
HPLC method	The product of nattokinase hydrolysis of peptide A (LKRLKRFLKRLK) was detected by a diode array detector and an Eclipse XDB-C18 column.	The operation process is complicated	[58]
Milk plate method	Nattokinase hydrolyzes casein on the plate to form a hydrolysis circle	Simple and fast, used for preliminary characterization of nattokinase activity	[59]
Serum plate method	Nattokinase hydrolyzes fibrin in serum to form hydrolysis circle	Low cost, convenient and quick; easily affected by the quality of the prepared plate.	[60]
Enzyme-linked immunosorbent method	Determination of nattokinase activity by specific enzyme-linked adsorption	High sensitivity and specificity; complicated operation and high cost	[61]
UV spectrophotometer method	Determination of the fibrinolytic activity of nattokinase by measuring the change in absorbance of nattokinase fibrin hydrolysate at 275 nm	The operation process is complicated	[62]