TY - JOUR
T1 - Purification and partial biochemical characterization of polyphenol oxidase from mamey (Pouteria sapota)
AU - Palma-Orozco, Gisela
AU - Ortiz-Moreno, Alicia
AU - Dorantes-Álvarez, Lidia
AU - Sampedro, José G.
AU - Nájera, Hugo
N1 - Funding Information:
We thank Dr. D. Alejandro Fernández-Velasco for facilities at the Laboratorio de Fisicoquímica e Ingeniería de Proteínas, UNAM. This work was partially supported by scholarship CONACyT ( 202061 ) GPO, and grants from IPN ( 20082319 , 20070660 , 20060529 , 20090612 ) AOM and LDA, CONACYT ( 105532 ) and PROMEP ( UAM-PTC-144 ) H.N.
PY - 2011/1
Y1 - 2011/1
N2 - While a long shelf life for fruit products is highly desired, enzymatic browning is the main cause of quality loss in fruits and is therefore a main problem for the food industry. In this study polyphenol oxidase (PPO), the main enzyme responsible for browning was isolated from mamey fruit (Pouteria sapota) and characterized biochemically. Two isoenzymes (PPO 1 and PPO 2) were obtained upon ammonium sulfate precipitation and hydrophobic and ion exchange chromatography; PPO 1 was purified up to 6.6-fold with 0.28% yield, while PPO 2 could not be characterized as enzyme activity was completely lost after 24 h of storage. PPO 1 molecular weight was estimated to be 16.1 and 18 kDa by gel filtration and SDS-PAGE, respectively, indicating that the native state of the PPO 1 is a monomer. The optimum pH for PPO 1 activity was 7. The PPO 1 was determined to be maximum thermally stable up to 35 °C. Kinetic constants for PPO 1 were Km = 44 mM and Km = 1.3 mM using catechol and pyrogallol as substrate, respectively. The best substrates for PPO 1 were pyrogallol, 4-methylcatechol and catechol, while ascorbic acid and sodium metabisulfite were the most effective inhibitors.
AB - While a long shelf life for fruit products is highly desired, enzymatic browning is the main cause of quality loss in fruits and is therefore a main problem for the food industry. In this study polyphenol oxidase (PPO), the main enzyme responsible for browning was isolated from mamey fruit (Pouteria sapota) and characterized biochemically. Two isoenzymes (PPO 1 and PPO 2) were obtained upon ammonium sulfate precipitation and hydrophobic and ion exchange chromatography; PPO 1 was purified up to 6.6-fold with 0.28% yield, while PPO 2 could not be characterized as enzyme activity was completely lost after 24 h of storage. PPO 1 molecular weight was estimated to be 16.1 and 18 kDa by gel filtration and SDS-PAGE, respectively, indicating that the native state of the PPO 1 is a monomer. The optimum pH for PPO 1 activity was 7. The PPO 1 was determined to be maximum thermally stable up to 35 °C. Kinetic constants for PPO 1 were Km = 44 mM and Km = 1.3 mM using catechol and pyrogallol as substrate, respectively. The best substrates for PPO 1 were pyrogallol, 4-methylcatechol and catechol, while ascorbic acid and sodium metabisulfite were the most effective inhibitors.
KW - Catechol
KW - PPO purification
KW - Polyphenol oxidase
KW - Pyrogallol
KW - Sapotaceae
KW - Sapote mamey (Pouteria sapota)
UR - http://www.scopus.com/inward/record.url?scp=78650712936&partnerID=8YFLogxK
U2 - 10.1016/j.phytochem.2010.10.011
DO - 10.1016/j.phytochem.2010.10.011
M3 - Artículo
C2 - 21087780
SN - 0031-9422
VL - 72
SP - 82
EP - 88
JO - Phytochemistry
JF - Phytochemistry
IS - 1
ER -