تولید و بهینه سازی نانو سامانه های کیتوزان برای حمل فراکسیون پپتیدی حاصل از هیدرولیز آنزیمی ضایعات سر میگوی وانامی (litopenaeus vannamei)

اﯾﻦ ﺗﺤﻘﯿﻖ ﺑﺎ ﻫﺪف ﺑﺮرﺳﯽ و ﺑﻬﯿﻨﻪﺳﺎزی ﮐﭙﺴﻮﻟﻪﺳﺎزی ﻓﺮاﮐﺴﯿﻮن ﭘﭙﺘﯿﺪی ﮐﻮﭼﮑﺘﺮ از 10 کلیودالتون ﺣﺎﺻﻞ از ﻫﯿﺪروﻟﯿﺰ آﻧﺰﯾﻤﯽ ﺿﺎﯾﻌﺎت ﺳﺮ ﻣﯿﮕﻮی واﻧﺎﻣﯽ (litopenaeus vannamei) ﺗﻮﺳط ﮐﯿﺘﻮزان اﻧﺠﺎم ﺷﺪ. ﻫﯿﺪروﻟﯿﺰ ﭘﺮوﺗﺌﯿﻨﯽ ﯾﺪ ﺳﺖ آﻣﺪه ﺗﻮ ﺳﻂ اوﻟﺘﺮاﻓﯿﻠﺘﺮا ﺳﯿﻮن ﺑﺎ اﻧﺪازه ﭼ ﺸﻤﻪ 10 ﮐﯿﻠﻮ داﻟﺘﻮن ﻓﺮاﮐﺴﯿﻮن ﺑﻨﺪی و ﻓﺮاﮐﺴﯿﻮن ﻣﻮرد ﻧﻈﺮ ﺑﺮ اﺳﺎس روش ژﻻﺳﯿﻮن ﯾﻮﻧﯽ (ﮐﯿﺘﻮزان-ﺗﺮی ﭘﻠﯽ ﺴﻔﺎت) در ﮐﭙﺴﻮل ﻫﺎی ﻧﺎﻧﻮﮐﯿﺘﻮزان ﮐﭙﺴﻮﻟﻪ و در ﺷﺮاﯾﻂ آزﻣﺎﯾﺸﮕﺎﻫﯽ ﺑﺮ اﺳﺎس ﻧﺴﺒﺖ ﻫﺎی ﻣﺨﺘﻠﻒ ﮐﯿﺘﻮزان:ﺗﺮی ﭘﻠﯽ ﻓﺴﻔﺎت و ﻏﻠﻈﺖ ﻫﺎی ﻣﺨﺘﻠﻒ ﻓﺮاﮐﺴﯿﻮن ﭘﭙﺘﯿﺪی ﺷﺎﻣﻞ 5 ،1 و 10 ﻣﯿﻠﯽ ﮔﺮم در ﻣﯿﻠﯽﻟﯿﺘﺮ ﺑﻬﯿﻨﻪ ﺳﺎزی ﺷﺪ. ﺑﺮ اﺳﺎس ﻧﺘﺎﯾﺞ ﻣﺸﺨﺺ ﺷﺪ ﮐﻪ آﻧﺰﯾﻢ آﻟﮑﺎﻻز ﺑﺎ درﺟﻪ ﻫﯿﺪروﻟﯿﺰ 57±2/37 درﺻﺪ ﺿﺎﯾﻌﺎت ﺳﺮ ﻣﯿﮕﻮ را ﻫﯿﺪروﻟﯿﺰ و ﻃﺒﻖ ﺷﺮاﯾﻂ اﯾﻦ آزﻣﺎﯾﺶ ﻃﻮل زﻧﺠﯿﺮه ﭘﭙﺘﯿﺪی 1/175 ﺣﺎﺻﻞ ﺷﺪ. وزن ﻣﻮﻟﮑﻮﻟﯽ ﭘﭙﺘﯿﺪ ﻫﺎی ﺣﺎﺿﺮ در ﻓﺮاﮐﺴﯿﻮن ﺣﺎﺻﻞ از اوﻟﺘﺮاﻓﯿﻠﺘﺮاﺳﯿﻮن دارای داﻣﻨﻪ وزﻧﯽ ﻧﺰدﯾﮏ ﺑﻬﻢ و زﯾﺮ 10 ﮐﯿﻠﻮ داﻟﺘﻮن ﺑﻮد. اﻧﺪازه ذرات در ﻏﻠﻈﺖﻫﺎ و ﺗﻤﯿﺎرﻫﺎی ﻣﺨﺘﻠﻒ ﻣﺘﻨﻮع و در ﻣﺤﺪوده 30 ﺗﺎ 150 ﻧﺎﻧﻮﻣﺘﺮ ﺑﻮد. ﺑﻬﺘﺮﯾﻦ ﻧﺘﯿﺠﻪ ﺣﺎ ﺻﻞ از ﺑﻬﯿﻨﻪ ﺳﺎزی ﮐﭙ ﺴﻮﻟﻪ ﺳﺎزی در ﺗﯿﻤﺎر ﻧ ﺴﺒﺖ 2:1 ﮐﯿﺘﻮزان ﺑﻪ ﭘﻠﯽ ﻓﺴﻔﺎت و ﻏﻠﻈﺖ 10 ﻣﯿﻠﯽ ﮔﺮم ﺑﺮ ﻣﯿﻠﯽﻟﯿﺘﺮ ﺑﺪﺳﺖ آﻣﺪ. اﻧﺪازه، ﺷﺎﺧﺺ ﭘﺮاﮐﻨﺪﮔﯽ (pdi)، ﭘﺘﺎﻧﺴﯿﻞ زﺗﺎ و اﻧﺪازه ﻧﺎﻧﻮﮐﭙﺴﻮل ﻫﺎ در ﺷﺮاﯾﻂ ﺑﻬﯿﻨﻪ ﻣﻮرد ﻧﻈﺮ ﺑﻪ ﺗﺮﺗﯿﺐ 2020 ،0/375 و 30/13 ﻧﺎﻧﻮﻣﺘﺮ ﺑﻮدﻧﺪ و ﺷﺮاﯾﻂ ﻧﮕﻬﺪاری در دﻣﺎی 20- درﺟﻪ ﺳﺎﻧﺘﯿﮕﺮاد ﺗﺎﺛﯿﺮ ﭼﻨﺪاﻧﯽ ﺑﺮ ﮐﯿﻔﯿﺖ ﻧﺎﻧﻮﮐﭙﺴﻮل ﻫﺎ ﻧﺪاﺷﺖ. ﯾﺎﻓﺘﻪ ﻫﺎی اﯾﻦ ﺗﺤﻘﯿﻖ ﻧﺸﺎن داد ﮐﯿﺘﻮزان ﻣﯽ ﺗﻮاﻧﺪ ﺑﺨﻮﺑﯽ ﺑﺮای ﻧﺎﻧﮑﭙ ﺴﻮﻟﻪ ﮐﺮدن ﻓﺮاﮐﺴﯿﻮن ﭘﭙﺘﯿﺪی ﮐﻮﭼﮑﺘﺮ از 10 ﮐﯿﻠﻮداﻟﺘﻮن ﺑﺎ ﮐﺎراﯾﯽ 91/04±0/18 درﺻﺪ ﻣﻮرد اﺳﺘﻔﺎده ﻗﺮار ﮔﯿﺮد.

Production and optimization of chitosan nanosystems loaded with peptide fraction resulting from enzymatic hydrolysis of vannamei shrimp (Litopenaeus vannamei) head wastes

In this experiment, head wastes were prepared and enzymatically hydrolyzed using alcalase (2.4 L) enzyme. The hydrolysate was fractionated by ultrafiltration with 10 kDa molecular weight cutoffs and the desired fraction was encapsulated following ion coagulation method (chitosan and triphosphate (TPP)) in nanochitosan capsules. Encapsulation process was optimized based on different ratios of chitosan:TPP and different concentrations (1, 5 and 10 mg/ml) of peptidic fraction. Finally, the degree of hydrolysis and the length of the peptides obtained from enzymatic hydrolysis were determined. The nanocapsules were examined for size, zeta potential and polydispersity index (PDI) using dynamic light scattering (Malvern, England). Structural and surface morphology studies including scanning electron microscopy (SEM) and infrared spectroscopy (FTIR) of capsules produced under favorable conditions were also performed. Particle size was measured in various concentrations and treatments in the range of 30 to 150 nm. The best results were obtained in the treatment of 2: 1 ratio of chitosan to polyphosphate and concentration of 10 mg / ml. The size, dispersion index, zeta potential and size of nanocapsules in the optimal conditions were 0.375, 2020 and 30.13 nm, respectively, and storage conditions at 20 °C had no effect on the quality of nanocapsules. Based on the efficiency study, it was found that fraction with a concentration of 10 mg/ml is well encapsulated by chitosan with an efficiency of 91.04 ± 0.18 percent. The results showed that chitosanTPP could be used for nanocapsulation of bioactive peptides with an approximate molecular weight of less than 10 kDa.