Restricted-access materials （RAMs） have found their broad application in sample pretreatment of bioanalysis. Boronate affinity （BA） adsorption is a very efficient and facile method for isolation and enrichment of ...Restricted-access materials （RAMs） have found their broad application in sample pretreatment of bioanalysis. Boronate affinity （BA） adsorption is a very efficient and facile method for isolation and enrichment of cis-diol containing biomolecules which are a large important group compounds in biosystems. However, preparation of BA-RAMs are rarely reported to date. In this study, a novel BA-RAM with external surface comprised of hydrophilic bottlebrush polymers was prepared exploiting the excellent capability of the bottlebrush polymers for protein exclusion. A diblock copolymer poly（3- acrylamidophenylboronic acid）-block-poly（2-hydroxyethyl methacrylate） （PAAPBA-b-PHEMA） was first grafted from the silica surface via surface-initiated reversible addition-fragmentation chain transfer polymerization （SI-RAFT）, and poly（N-isopropylacrylamide） （PNIPAAm） was then grafted from the PHEMA via surface-initiated atom transfer radical polymerization （SI-ATRP） to yield the BA-RAM. The BA- RAM exhibits high selectivity to cis-diol containing small molecules and has good capability to exclude proteins. Its practical application in bioanalysis was tested by pretreatment of serum sample for analysis of catecholamines with high recoveries and good precision. The preparation strategy for the BA-RAM is very versatile and is easy to be expanded to other modes of RAMs.展开更多
The adsorption and separation of antibody drugs are of great significance,but the promising hydrophobic charge induction chromatography(HCIC)and boronate affinity chromatography(BAC)suffer from low specific due to the...The adsorption and separation of antibody drugs are of great significance,but the promising hydrophobic charge induction chromatography(HCIC)and boronate affinity chromatography(BAC)suffer from low specific due to the limitations of single-site adsorption mechanism as well as low adsorption capacity of adsorbents,resulting in a lower purity and recovery of antibodies.To address this issue,this work proposes a two-site synergistic binding strategy integrating HCIC and BAC mechanism on a polymer brushes-grafted adsorbent.Five adsorbents were easily created by polymerizing the mixed monomers of 5-acryloylaminobenzimidazole,3-acryloylamide phenylboronic acid and acrylamide on surface of agarose gel via activators regenerated by electron transfer for atom transfer radical polymerization(ARGET ATRP).The molecular docking implies that the two-site synergistic binding towards immunoglobulin G(IgG)originates from the closely adjacent boronic and benzimidazole side groups in the polymer chains with monomer ratio of 1:1:0.The inference was verified by the effect of three monomer ratios and adsorption conditions on the adsorption performance of IgG.The adsorbent with two-site synergy possesses an excellent specific,enhanced affinity(K_(d)=3.9×10^(-6)mol/L)and adsorption capacity(Q_(m)=253 mg/g)towards IgG.Benefiting from the advantages,IgG from serum and monoclonal antibody(m Ab)from cell culture achieve purities of 95.8%and 98.3%,and recoveries of 95.7%and 97.5%,respectively.The results are comparable to those with protein A adsorbent considered to have the best specific so far,indicating the potential of the two-site synergistic binding strategy in the purification of antibody drugs.展开更多
20-Hydroxyecdysone(20E)derived from plants has a wide range of physiological and pharmacological ef ects on animals and humans,and rapid and sensitive methods for screening of the endogenous 20E in plants are thus req...20-Hydroxyecdysone(20E)derived from plants has a wide range of physiological and pharmacological ef ects on animals and humans,and rapid and sensitive methods for screening of the endogenous 20E in plants are thus required.Herein,a matrixassisted laser desorption/ionization time-of-f ight tandem mass spectrometry(MALDI-TOF/TOF MS)method is described for rapid and sensitive determination of endogenous 20E in plants.It is based on the use of the(3-(acridin-9-ylamino)phenyl)boronic acid(AYPBA)as the mass tag to assist the MS and tandem MS(MS^(n))analysis of 20E on MALDI-TOF/TOF MS.Good linearity was obtained with a determination coef cient(R^(2))larger than 0.99 in the range of 0.025–2.5μΜ.The limit of detection(LOD)was 2.4 fmol.Acceptable precision and accuracy were gained by intra-and inter-day analysis with relative standard deviations less than 19.5%and relative recoveries ranging from 85.7 to 105.2%.In addition,the AYPBA labeled 20E produced abundant characteristic fragment ions under the high energy collision-induced dissociation,which facilitated the identif cation of 20E by MS^(2)analysis on MALDI-TOF/TOF MS.Using the method,we enabled the identif cation and quantif cation of endogenous 20E in four herbs including Cyanotis arachoidea,Achyranthes bidentata,Spinacia oleracea and Chenopodium quinoa willd.,demonstrating the feasibility of the proposed method for screening of the endogenous 20E in plants.展开更多
基金supported by the National Natural Science Foundation of China(Nos. 21575114 and 21475104)the Scientific Research Program Funded by Shaanxi Provincial Education Department(No. 16JS114)
文摘Restricted-access materials （RAMs） have found their broad application in sample pretreatment of bioanalysis. Boronate affinity （BA） adsorption is a very efficient and facile method for isolation and enrichment of cis-diol containing biomolecules which are a large important group compounds in biosystems. However, preparation of BA-RAMs are rarely reported to date. In this study, a novel BA-RAM with external surface comprised of hydrophilic bottlebrush polymers was prepared exploiting the excellent capability of the bottlebrush polymers for protein exclusion. A diblock copolymer poly（3- acrylamidophenylboronic acid）-block-poly（2-hydroxyethyl methacrylate） （PAAPBA-b-PHEMA） was first grafted from the silica surface via surface-initiated reversible addition-fragmentation chain transfer polymerization （SI-RAFT）, and poly（N-isopropylacrylamide） （PNIPAAm） was then grafted from the PHEMA via surface-initiated atom transfer radical polymerization （SI-ATRP） to yield the BA-RAM. The BA- RAM exhibits high selectivity to cis-diol containing small molecules and has good capability to exclude proteins. Its practical application in bioanalysis was tested by pretreatment of serum sample for analysis of catecholamines with high recoveries and good precision. The preparation strategy for the BA-RAM is very versatile and is easy to be expanded to other modes of RAMs.
基金supported by the National Natural Science Foundation of China(Nos.22274129,21974106 and 22074117)。
文摘The adsorption and separation of antibody drugs are of great significance,but the promising hydrophobic charge induction chromatography(HCIC)and boronate affinity chromatography(BAC)suffer from low specific due to the limitations of single-site adsorption mechanism as well as low adsorption capacity of adsorbents,resulting in a lower purity and recovery of antibodies.To address this issue,this work proposes a two-site synergistic binding strategy integrating HCIC and BAC mechanism on a polymer brushes-grafted adsorbent.Five adsorbents were easily created by polymerizing the mixed monomers of 5-acryloylaminobenzimidazole,3-acryloylamide phenylboronic acid and acrylamide on surface of agarose gel via activators regenerated by electron transfer for atom transfer radical polymerization(ARGET ATRP).The molecular docking implies that the two-site synergistic binding towards immunoglobulin G(IgG)originates from the closely adjacent boronic and benzimidazole side groups in the polymer chains with monomer ratio of 1:1:0.The inference was verified by the effect of three monomer ratios and adsorption conditions on the adsorption performance of IgG.The adsorbent with two-site synergy possesses an excellent specific,enhanced affinity(K_(d)=3.9×10^(-6)mol/L)and adsorption capacity(Q_(m)=253 mg/g)towards IgG.Benefiting from the advantages,IgG from serum and monoclonal antibody(m Ab)from cell culture achieve purities of 95.8%and 98.3%,and recoveries of 95.7%and 97.5%,respectively.The results are comparable to those with protein A adsorbent considered to have the best specific so far,indicating the potential of the two-site synergistic binding strategy in the purification of antibody drugs.
基金supported by the National Key R&D Program of China(2017YFC0906800)the National Natural Science Foundation of China(21635006,31670373 and 21721005)
文摘20-Hydroxyecdysone(20E)derived from plants has a wide range of physiological and pharmacological ef ects on animals and humans,and rapid and sensitive methods for screening of the endogenous 20E in plants are thus required.Herein,a matrixassisted laser desorption/ionization time-of-f ight tandem mass spectrometry(MALDI-TOF/TOF MS)method is described for rapid and sensitive determination of endogenous 20E in plants.It is based on the use of the(3-(acridin-9-ylamino)phenyl)boronic acid(AYPBA)as the mass tag to assist the MS and tandem MS(MS^(n))analysis of 20E on MALDI-TOF/TOF MS.Good linearity was obtained with a determination coef cient(R^(2))larger than 0.99 in the range of 0.025–2.5μΜ.The limit of detection(LOD)was 2.4 fmol.Acceptable precision and accuracy were gained by intra-and inter-day analysis with relative standard deviations less than 19.5%and relative recoveries ranging from 85.7 to 105.2%.In addition,the AYPBA labeled 20E produced abundant characteristic fragment ions under the high energy collision-induced dissociation,which facilitated the identif cation of 20E by MS^(2)analysis on MALDI-TOF/TOF MS.Using the method,we enabled the identif cation and quantif cation of endogenous 20E in four herbs including Cyanotis arachoidea,Achyranthes bidentata,Spinacia oleracea and Chenopodium quinoa willd.,demonstrating the feasibility of the proposed method for screening of the endogenous 20E in plants.
