The prote
ctive effe
cts of amino a
cids on stabilizing protein se
condary stru
cture were evaluated using diffuse refle
ctan
ce FTIR spe
ctros
copy, and intera
ctions between proteins and arginine were dete
cted using solid-state NMR spe
ctros
copy. Upon freeze-drying, ex
cipient-free anti-CD11a and anti-IgE antibodies underwent signifi
cant
changes in their se
condary stru
ctures. For both antibodies, the amount of intermole
cular β-sheet substantially in
creased and the native
conformation of intramole
cular β-sheet
content de
creased
considerably. The addition of amino a
cids to the formulations redu
ced protein se
condary stru
cture alterations in a
con
centration-dependent manner. Histidine and arginine appeared to be the most prote
ctive ex
cipients (of the amino a
cids studied) in inhibiting protein se
condary stru
ctural
changes. Solid-state NMR illustrated that non-
covalent intera
ctions (e.g., hydrogen bonding, ion–dipole intera
ctions) were formed between the arginine side
chain and the protein. Gly
cine is the least effe
ctive additive of those studied in preventing se
condary stru
cture
changes upon freeze-drying.
Despite secondary structural changes, freeze-dried protein in the presence and absence of amino acids refolded back into its native conformation upon reconstitution in water.