Study On The Browning Mechanism Of Betel Nut Kernel

The storage process of betel nut(Areca catechu L.)is highly prone to browning of the kernel and affects the quality of the fruit.In this study,betel nut was used as the raw material.Firstly,the key conditions of enzymatic browning in fresh betel nut,phenols and polyphenol oxidase(PPO),were extracted and purified to determine the possibility of enzymatic browning;further studies on browning products were carried out to clarify the relationship between betel nut browning and enzymatic browning.Then,the incentives of betel nut kernel browning was studied from the temperature point of view.On this basis,the focus is on the mechanism of browning of kernel from three aspects:the relationship between the change of phenolic content,the activity browning-related enzymes and browning of areca nut kernel,and how the cell regionalization is broken,leading to the contact between PPO and phenolic substances.And how the O₂ component enters the inside of the kernel,so that the mechanism of browning of the kernel during storage of fresh betel nut is clearly understood.The research results are as follows:(1)11 kinds of phenolic compounds in betel nut kernel were separated by liquid-mass spectrometry,among which chlorogenic acid was the highest,followed by dopamine and L-epicatechin,and other contents were relatively low.At the same time,PPO was also isolated from the betel nut kernel,the relative molecular mass was 29.2 kDa,the optimum temperature of PPO in betel nut kernel was 20?,and the optimum pH was 7.0.With catechol as substrate,the maximum reaction rate(Vmax)of the enzyme was 140.84 U/mL min and Km was 3.22 mmol/L.On the basis of this,the spectral comparison of the oxidation product obtained by the separation and purification of PPO with chlorogenic acid,dopamine and epicatechin in vitro and the product extracted from brown betel nut showed that the two had the same absorption spectrum.Therefore,it is speculated that the browning of betel nut kernel is an enzymatic browning caused by the reaction of PPO and phenolic substrates.(2)By studying the degree of browning of betel nut kernel in postharvest storage at 5,10,15 and 25?,the results showed that the storage temperature significantly affected the browning degree of betel nut kernel,between 10-25 C,with the increase of temperature,fruit ripening and aging accelerated,and browning of betel nut kernel increased.With the occurrence of chilling injury at 5?,the browning of betel nut kernel was also aggravated,but the browning degree is lower than the betel nut fruit at high temperature,indicating that chilling damage is not the only factor that causes browning of betel nut kernel.Both chilling and high temperature mature senescence can induce browning of betel nut kernel.And the faster the total phenolic content decreases,the higher the PPO and PAL activities of the browning-related enzymes,the more serious the browning.(3)The higher the PPO and PAL activities of the browning-related enzymes,the more the content of phenolic substances in the enzymatic browning substrate decreased,and the browning degree of the kernel was more serious of betel nut stored at different temperatures.It is further explained that betel nut kernel browning is associated with enzymatic browning.(4)During storage,the catalase(CAT)activity of betel nut kernel increased with increasing temperature in the range of 10-25?,while the CAT activity under chilling was only lower than that of betel nut kernel at 25?.The superoxide dismutase(SOD)activity increases with increasing temperature,whereas in contrast,peroxidase(POD)activity increases with decreasing temperature.However,the active oxygen scavenging enzymes of SOD,CAT and POD could not completely remove excess reactive oxygen species,so the temperature rise and chilling could cause the H₂O₂ and O₂^-· content in the betel nut kernel to rise rapidly,causing the membrane lipid peroxidation product.The MDA increases,which causes the membrane lipid peroxidation product MDA to increase,the membrane permeability increases,the cell membrane tissue structure is destroyed,resulting in destruction of tissue regionalization.(5)Scanning electron microscopy showed that the waxy distribution of betel nut surface was uniform and dense in the initial stage of storage,and the structure was intact without cracks.The flesh tissue was arranged tightly and without gaps,and the surface of the kernel membrane was smooth and wrinkle-free.With the prolonged storage time,the temperature rising and chilling will lead to the damage of fruit tissue.The damage of the tissue at 25?is the most serious,the cuticle of the peel is honeycomb-shaped,the wax is gradually decomposed;the lignification of the flesh is aggravated,the tissue gap increases;the kernel membrane cracks and breaks.Secondly,there is a hole in the keratin membrane under chilling at 5?,the waxy coverage is reduced,the flesh tissue is gradually loosened and separated,and the kernel membrane is cracked and picked up.These changes help to increase the exchange of gases inside and outside the organization,including the entry of oxygen.In summary,both chilling injury and high temperature can induce browning of betel nut kernel.As the temperature rises and chilling occurs,PPO activity increases,and the accumulation of reactive oxygen species causes membrane structure damage and cell regionalization is destroyed.The key enzyme PPO which causes enzymatic browning is in contact with the phenolic substrate.At the same time,with the increase of storage time,chilling damage and high temperature increase the damage of the betel nut peel,flesh and kernel membrane structure,O₂ penetrates into the kernel,as a result,phenolic substrates were oxidized by PPO to induce enzymatic browning of kernel.