HPLC analysis of functionalized poly(amidoamine) dendrimers and the interaction between a folate-dendrimer conjugate and folate binding protein.
Poly(amidoamine) (PAMAM) dendrimers of different generations with carboxyl, acetyl, and hydroxyl terminal groups and a folic acid (FA)-dendrimer conjugate were separated and analyzed using reverse-phase high performance liquid chromatography (HPLC). Analysis of both the individual PAMAM derivatives and the separation of mixed generations can be achieved using a linear gradient 0-50% acetonitrile (ACN) (balance water) within 40 min. We also show that PAMAMs with defined acetylation and carboxylation degrees can be analyzed using HPLC. Furthermore, a generation 5 dendrimer-FA conjugate (G5.75Ac-FA4; Ac denotes acetyl) was analyzed and its specific binding with a bovine folic acid binding protein (FBP) was monitored. The HPLC and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) results indicate the formation of three complexes after the binding of G5.75Ac-FA4 with FBP. Dendrimers with FA moieties show much higher specific binding capability with FBP than those without FA moieties. Findings from this study indicate that HPLC is an effective technique not only for characterization and separation of functionalized PAMAM dendrimers and conjugates but also for investigation of the interaction between dendrimers and biomolecules.
Shi
Xiangyang
X
Michigan Nanotechnology Institute for Medicine and Biological Sciences, University of Michigan, Ann Arbor, MI 48109, USA.
Bi
Xiangdong
X
Ganser
T Rose
TR
Hong
Seungpyo
S
Myc
Lukasz A
LA
Desai
Ankur
A
Holl
Mark M Banaszak
MM
Baker
James R
JR
D001672:Biocompatible Materials / Q000032:analysis / Q000737:chemistry; D002352:Carrier Proteins / Q000737:chemistry*; D002850:Chromatography, Gel / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D050091:Dendrimers; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D058973:Folate Receptors, GPI-Anchored; D011073:Polyamines / Q000032:analysis* / Q000737:chemistry; D011956:Receptors, Cell Surface / Q000737:chemistry*
D001672:Biocompatible Materials; D002352:Carrier Proteins; D050091:Dendrimers; D058973:Folate Receptors, GPI-Anchored; C104700:PAMAM Starburst; D011073:Polyamines; D011956:Receptors, Cell Surface