Application And Analysis Of Deep Eutectic Solvent Based Aqueous Bisphasic Systems For Partitioning Biomacromolecules

Owing to the ease of preparation and the convenience of utilization,deep eutectic solvent?DES?has been applied in various field s,especially in aqueous biphasic systems?ABSs?.Derived from two water-soluble compounds,ABSs provide many advantages,involving sustainable and biocompatible purification environment.According to numerous exploration,it is found that DES-based ABSs not only exhibit high extraction efficiency,but also protect the integrity of analytes.Besides,the systems also have the facility to enormously prepare in terms of industrialization.It is of importance for proteins and nucleic acid to be separated and further purified because they play a significant role in living activities.Many pigments has been widely applied in the food and spinning industry.And it is an paramount measure to accurately and quickly detect the forbidden pigments with the intention of ensuring food safety.In this work,many kinds of novel deep eutectic solvents were successfully synthesized.A couple of new-type salts were explored for their salting-out ability.Combined the prepared deep eutectic solvents with salts,ABSs were estab lished for high-performance partitioning of bovine serum albumin?BSA?,RNA and amaranth.The main work in detail is shown as follows:?1?Ternary and binary deep eutectic solvents as a novel extraction media for protein partitioningThis work was devoted to making a comparison among ternary and binary deep eutectic solvents with aqueous two-phase system to extract proteins.Four kinds of binary DESs and four ternary DESs which comprising of hydrogen bond acceptor?etramethylammonium chloride,TMAC?and hydrogen bond donors,were synthesized.The phase diagrams of the studied DESs demonstrated that binary DESs were more amenable to produce a two-phase splitting.Afterwards,the ability to extract four different substances was further evaluated.It was found that the systems based on ternary DESs showed a better potential in the field of extraction.Two kinds of DESs with ATPS,involving TMAC-urea?TMAC-U?and TMAC-glycerol-urea?TMAC-G-U?,were investigated among five influence factors such as the amount of DES,the concentration of the salt,the mass of BSA,the temperature and the pH value in terms of extracting bovine serum albumin?BSA?.Under the optimal conditions,the values of their respective extraction efficiency were up to 99.31%and 98.95%.Moreover,the back extraction efficiency were discussed and indicated that the ternary DESs reached to 71.89%,which was remarkably different with binary DES?21.02%?.Finally,UV-vis,fourier transform infrared spectrometry?FT-IR?,circular dichroism?CD?,dynamic light scattering?DLS?and transmission electron microscopy?TEM?were utilized to study the extraction mechanism.This work reveals the outstanding feasibility of TMAC-G-U based DES with ATPS in the protein extraction process,providing significant m ethod for designing task-specific DESs.?2?Aqueous biphasic systems containing PEG-based deep eutectic solvents for high-performance partitioning of RNAIn this work,16 kinds of novel deep eutectic solvents composed of polyethylene glycol?PEG?and quaternary ammonium salts,were coupled with Aqueous Biphasic Systems to extract RNA.The phase forming ability of ABSs were comprehensively evaluated,involving the effects of various proportions of DESs'components,carbon chain length and anions species of quaternary ammonium salts,average molecular weights of PEG and inorganic salts nature.Then the systems were applied in RNA extraction,and the results revealed that the extraction efficiency values were distinctly enhanced by relatively lower PEG content in DESs,smaller PEG molecular weights,longer carbon chain of quaternary ammonium salts and more hydrophobic inorganic salts.Then the systems composed of[TBAB][PEG600]and Na₂SO₄ were utilized in the influence factor experiments,proving that electrosta tic interaction was the dominant force for RNA extraction.Therefore,back-extraction efficiency values ranging between 85.19%and 90.78%were obtained by adjusting the ionic strength.Besides,the selective separation of RNA and tryptophane?Trp?was successfully accomplished.It was found that 86.19%RNA was distributed in the bottom phase,while 72.02%Trp was enriched in the top phase in the novel ABSs.Finally,dynamic light scattering and transmission electron microscope were used to further investigate the extraction mechanism.The proposed method reveals the outstanding feasibility of the newly developed ABSs formed by PEG-based DESs and inorganic salts for the green extraction of RNA.?3?Aqueous biphasic systems formed by PPG-based deep eutectic solvent for extraction of amaranthIn this work,deep eutectic solvent composed of polypropylene glycol 400?PPG400?and tetrabutylammonium bromide?TBAB?was combined with a series of new-type salts to form Aqueous Biphasic Systems,such as quaternary ammo nium salts,amino acid and polyols.Phase-forming ability of the salts was investigated firstly.It was clear that polyols had a relatively weak power to produce phases within studied scopes.And the shorter of carbon chain length of salts,the easier to obtain phase-splitting.Then partitioning of three pigments in PPG 400/betaine-based ABSs was addressed to investigate the effect of hydrophobicity on extraction efficiency.It was found that an increase in hydrophobicity contributed to the migration of pigments in the DES-rich phase.On the other hand,with a decline in phase-forming ability of salts,the extraction efficiency of the whole systems started to go down gradually.Based on the results,selective separation experiment was conducted successfully in the PPG 400/betaine-based systems,including more than93.00%Sudan?in the top phase and about 80.00%sunset yellow FCF/amaranth in the bottom phase.Additionally,ABSs constructed by DES/betaine for partitioning amaranth were further utilized to explore the performances of influence factors and back extraction.It can be concluded that after the optimization above 98.00%amaranth was transferred into the top phase.And 67.98%amaranth can be transferred into the bottom phase in back-extraction experiment.At last,DLS and TEM were applied to probe into extraction mechanism.It was demonstrated that hydrophobicity played an important role in the separation process of pigments.This work was devoted to introducing plentiful salts as novel compositions of ABSs,and providing an eco-friendly extraction way for partitioning pigments which boosted development of ABSs in the monitoring food safety field.