title: FTIR and nDSC as analytical tools for high-concentration protein formulations.
abstract: OBJECTIVE
  The aim of the study is to evaluate Fourier-transform infrared spectroscopy (FTIR) as an analytical tool for high-concentrated protein formulations.
  
  
  METHODS
  FTIR is used to determine the melting temperature (T(m (FTIR))) of various proteins, such as bovine serum albumin (BSA), immunoglobulin (IgG1), beta-lactoglobulin (beta-LG), and lysozyme (HEWL), at different protein concentrations (5-100 mg/mL), where four data interpretation methods are discussed. The obtained T(m (FTIR)) values are further compared to the T(m) measured by the nanodifferential scanning calorimetry (nDSC) technique.
  
  
  RESULTS
  The T(m (FTIR)) values of IgG1 and beta-LG showed strong consistency and corresponded to the nDSC results irrespective of the method of data interpretation and the protein concentration applied. In contrast, the T(m (FTIR)) of BSA and HEWL is characterized by significant deviations. Only the midpoint of the second-derivative intensity-temperature curve of the intermolecular beta-sheet mode measured at a concentration of 100 mg/mL is consistent with the nDSC results.
  
  
  CONCLUSIONS
  Determination of a T(m (FTIR)) is feasible by the midpoint of the intensity-temperature plot of the arising intermolecular beta-sheet band. More significant results are obtained for proteins, which are predominantly composed of intramolecular beta-sheet elements as well as at higher protein concentrations. A further study was started to assess the predictability of long-term protein stability by T(m (FTIR)).
authors.affiliation: Merck KGaA, Global Pharmaceutical Development, Darmstadt, Germany.
authors.forename: Susanne
authors.initials: S
authors.lastname: Matheus
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Friess
authors.forename: Hanns-Christian
authors.initials: HC
authors.lastname: Mahler
mesh_terms: D002152:Calorimetry, Differential Scanning; D002626:Chemistry, Pharmaceutical; D004795:Enzyme Stability; D007074:Immunoglobulin G / Q000737:chemistry*; D007782:Lactoglobulins / Q000737:chemistry*; D009113:Muramidase / Q000737:chemistry*; D036103:Nanotechnology; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared; D013816:Thermodynamics; D044366:Transition Temperature
chemical_list: D007074:Immunoglobulin G; D007782:Lactoglobulins; D012710:Serum Albumin, Bovine; D009113:Muramidase