تأثير فرآيندهاي انجماد- انجمادزدايي و خشك كردن انجمادي- رهيدراسيون بر پايداري فيزيكي نانوليپوزوم هاي حامل پپتيدهاي زيست فعال

The Effect of Freezing/thawing and Freeze-drying / Rehydration Processeson the Physical Stability of Nano-Liposomes Containing the Bioactive Peptides

در ﺳﺎلﻫﺎي اﺧﯿﺮ ﺿﺎﯾﻌﺎت ﺣﺎﺻﻞ از آﺑﺰﯾﺎن ﺑﻪ ﻋﻨﻮان ﻣﻨﺒﻌﯽ از ﭘﭙﺘﯿﺪﻫﺎي زﯾﺴﺖ ﻓﻌﺎل ﻣﻮرد ﺗﻮﺟﻪ ﺑﺴﯿﺎري از ﻣﺤﻘﻘﺎن ﻗﺮار ﮔﺮﻓﺘﻪاﺳﺖ. ﻟﯿﮑﻦ ﺗﻮﻟﯿﺪ ﭘﭙﺘﯿﺪﻫﺎي زﯾﺴﺖ ﻓﻌﺎل ﻧﯿﺎزﻣﻨﺪ روشﻫﺎي ﻣﺤﺎﻓﻈﺘﯽ ﻣﯽﺑﺎﺷﺪ ﺗﺎ ﻋﻼوه ﺑﺮ اﻓﺰاﯾﺶ ﭘﺎﯾﺪاري ﻓﯿﺰﯾﮑﯽ، ﻣﻨﺠﺮﺑﻪ رﻫﺎﯾﺶ ﮐﻨﺘﺮل ﺷﺪه و ﺑﻬﯿﻨﻪﺳﺎزي ﺗﺎﺛﯿﺮ آن ﻃﯽ ﻣﺼﺮف ﮔﺮدد. ﻟﺬا، ﺗﺤﻘﯿﻖ ﺣﺎﺿﺮ ﺑﺎ ﻫﺪف ﺗﻮﻟﯿﺪﻧﺎﻧﻮﻟﯿﭙﻮزومﻫﺎي ﺣﺎﻣﻞ ﭘﭙﺘﯿﺪﻫﺎي زﯾﺴﺖ ﻓﻌﺎل ﻣﺴﺘﺨﺮج از ﻣﺎﻫﯽ ﮐﭙﻮر ﻣﻌﻤﻮﻟﯽ )Cyprinus carpio( ﺑﺎ اﺟﺰاي ﭘﭙﺘﯿﺪي در ﻣﺤﺪوده زﯾﺮ 3 ﺗﺎ ﺑﺎﻻي 30 ﮐﯿﻠﻮداﻟﺘﻮن ﺑﻪ ﻣﻨﻈﻮر اﻓﺰاﯾﺶ ﭘﺎﯾﺪاري ﻓﯿﺰﯾﮑﯽ،ﺑﺎ ﭘﻮﺷﺶ ﮐﯿﺘﻮزان اﻧﺠﺎم ﮔﺮﻓﺖ.ﻧﺘﺎﯾﺞ ﻧﺸﺎن دادﻣﯿﺎﻧﮕﯿﻦ اﻧﺪازه ذرات ﻧﺎﻧﻮﻟﯿﭙﻮزومﻫﺎ در ﻣﺤﺪوده 333/9 ﺗﺎ 533/9 ﻧﺎﻧﻮﻣﺘﺮ ﺑﺎ ﭘﺘﺎﻧﺴﯿﻞ زﺗﺎي 54/03- ﺗﺎ46/3+ ﻣﯿﻠﯽ وﻟﺖ و ﺷﺎﺧﺺ ﭘﺮاﮐﻨﺪﮔﯽ )PDI( 0/2 ﺗﺎ 0/54 ﻗﺮار دارد. ﻣﻘﺎدﯾﺮ راﻧﺪﻣﺎن ﻧﺎﻧﻮﭘﻮﺷﺎﻧﯽ ﺑﻪ ﻃﻮر ﻗﺎﺑﻞ ﺗﻮﺟﻬﯽ ﻣﻮﺛﺮ از ﺗﻐﯿﯿﺮات ﻏﻠﻈﺖ ﮐﯿﺘﻮزان در ﭘﻮﺷﺶ ﺑﻮد و ﺣﺪﮐﺜﺮ راﻧﺪﻣﺎن ﻧﺎﻧﻮﭘﻮﺷﺎﻧﯽ )0/51 ± %72/57( در ﻧﺎﻧﻮﻟﯿﭙﻮزوم ﺣﺎﻣﻞ ﭘﭙﺘﯿﺪ ﺑﺎ وزن ﻣﻮﻟﮑﻮﻟﯽ ﮐﻤﺘﺮ از 3 ﮐﯿﻠﻮ داﻟﺘﻮن ﺑﺎ ﭘﻮﺷﺶ 1درﺻﺪ ﮐﯿﺘﻮزان ﻣﺸﺎﻫﺪه ﺷﺪ. ﺑﺮرﺳﯽ ﻣﯿﺰان رﻫﺎﯾﺶ ﭘﭙﺘﯿﺪ در ﻣﺤﯿﻂ ﺑﯿﺮون ﺗﻨﯽ ﺣﺎﮐﯽ از ﺗﺎﺛﯿﺮ ﭘﻮﺷﺶ ﺑﺮ ﮐﻨﺪ ﻧﻤﻮدن رﻫﺎﯾﺶ و اﻓﺰاﯾﺶ ﭘﺎﯾﺪاري ﭘﭙﺘﯿﺪﻫﺎ ﺑﻮد. ﺑﺎارزﯾﺎﺑﯽ ﭘﺎﯾﺪاري ﻓﯿﺰﯾﮑﯽ اﻧﺘﺸﺎر ﺳﺮﯾﻊﺗﺮ ﭘﭙﺘﯿﺪ از ﻟﯿﭙﻮزومﻫﺎ ﭘﺲ از ﻓﺮاﯾﻨﺪﻫﺎي اﻧﺠﻤﺎد/ اﻧﺠﻤﺎدزداﯾﯽ و ﺧﺸﮏﮐﺮدن اﻧﺠﻤﺎدي/ رﻫﯿﺪراﺳﯿﻮن ﻣﺸﺎﻫﺪه ﺷﺪ. ﻧﺘﺎﯾﺞ ﻧﮕﻬﺪاري دراز ﻣﺪت ﻧﺎﻧﻮﻟﯿﭙﻮزومﻫﺎ در دو دﻣﺎي 4 و 20 درﺟﻪ ﺳﺎﻧﺘﯽﮔﺮاد ﻣﺸﺨﺺ ﻧﻤﻮد ﭘﻮﺷﺶ ﮐﯿﺘﻮزان ﺑﺎ اﻓﺰودن ﺿﺨﺎﻣﺖ ﻏﺸﺎ و اﻓﺰاﯾﺶ داﻓﻌﻪ ﺑﯿﻦ ذرات ﺳﺒﺐ ﭘﺎﯾﺪاري ﺑﯿﺸﺘﺮ ﻧﺎﻧﻮﻟﯿﭙﻮزومﻫﺎ ﮔﺮدﯾﺪ ﻧﺘﺎﯾﺞ ﺗﺤﻘﯿﻖ ﺣﺎﺿﺮ ﻧﺸﺎن داد ﻧﺎﻧﻮﭘﻮﺷﺎﻧﯽ ﭘﭙﺘﯿﺪﻫﺎي زﯾﺴﺖ ﻓﻌﺎل در ﺣﺎﻣﻞ ﻟﯿﭙﻮزوﻣﯽ ﻣﯽﺗﻮاﻧﺪ روﺷﯽ ﻣﻨﺎﺳﺐ در ﺟﻬﺖ ﮐﺎرﺑﺮد در ﺻﻨﻌﺖ ﻏﺬا و دارو ﺑﺎﺷﺪ.

In recent years, the aquatic waste as a source of bioactive peptides has been considered by many researchers. Bioactive peptides require protective methods to increase physical stability and, controlled release and optimize efficacy during oral delivery. The aim of this study was to develop an oral phospholipid nanoliposomal system incorporated with bioactive peptides derived from fish protein hydrolysate of common carp (Cyprinus carpio) with separating fractions of peptide in range of 3 kDa to 30 kDa in chitosan coating (0.5%, 1% w/v) by the alkalase enzyme. The results showed that chitosan coating greatly improved the stability of nano-liposomes. The average particle size was in the range of 333.9- 533.9 nm with a zeta potential of -54.03 to +46.3 and a poly dispersity index (PDI) of 0.2- 0.54 in nano-liposomes. The Encapsulation Efficiency (EE%) values were significantly influenced by changes in the concentration of chitosan and the maximum EE% (72.57± 0.51) was observed in the nano-liposome containing peptide with 3 kDa fractions coated with 1% chitosan. Study on the invitro releasing rate of the peptide showed the effect of the coating on the slower release and peptide stability in nano-liposomes.Physical stability evaluation determined a faster release of the peptide from the liposomes after freezing/thawing and freeze-drying / rehydration processes. The results of long-term storage of nano-liposomes at temperatures of 4 and 20 ° C indicated that chitosan coating increased the stability of nano-liposomes by adding membrane thickness and increasing repulsion between the particles. The results of this study showed that encapsulation of bioactive peptide in the liposomal system could be a useful approach for direct application of peptides with bioactive potential in food and drug industry.