The application of STEP-technology® for particle and protein dispersion detection studies in biopharmaceutical research.

Particle detection and analysis techniques are essential in biopharmaceutical industries to evaluate the quality of various parenteral formulations regarding product safety, product quality and to meet the regulations set by the authority agencies. Several particle analysis systems are available on the market, but for the operator, it is quite challenging to identify the suitable method to analyze the sample. At the same time these techniques are the basis to gain a better understanding in biophysical processes, e.g. protein interaction and aggregation processes. The STEP-Technology® (Space and Time resolved Extinction Profiles), as used in the analytical photocentrifuge LUMiSizer®, has been shown to be an effective and promising technique to investigate particle suspensions and emulsions in various fields. In this study, we evaluated the potentials and limitations of this technique for biopharmaceutical model samples. For a first experimental approach, we measured silica and polystyrene (PS) particle standard suspensions with given particle density and refractive index (RI). The concluding evaluation was performed using a variety of relevant data sets to demonstrate the significant influences of the particle density for the final particle size distribution (PSD). The most challenging property required for successful detection, turbidity, was stated and limits have been set based on the depicted absorbance value at 320 nm (A320 values). Furthermore, we produced chemically cross-linked protein particle suspensions to model physically "stable" protein aggregates. These results of LUMiSizer® analysis have been compared to the orthogonal methods of nanoparticle tracking analysis (NTA), dynamic light scattering (DLS) and micro-flow imaging (MFI). Sedimentation velocity distributions showed similar tendencies, but the PSDs and absolute size values could not be obtained. In conclusion, we could demonstrate some applications as well as limitations of this technique for biopharmaceutical samples. In comparison to orthogonal methods this technique is a great complementary approach if particle data e.g. density or refractive index can be determined.

Relationship of prolonged pharmacologic serum levels of vitamin E to incidence of sepsis and necrotizing enterocolitis in infants with birth weight 1,500 grams or less.

The incidence of culture-proven neonatal sepsis and necrotizing enterocolitis (NEC) in preterm infants maintained at pharmacologic (mean 5.1 mg/dL +/- 1.45 SD) serum vitamin E levels for long periods was prospectively studied as part of a double-masked clinical trial of the effect of prophylactic vitamin E v placebo treatment on the development and course of retinopathy of prematurity (ROP). Within a few days of birth, 914 preterm infants were enrolled in the study; 545 (275 placebo-treated infants, 270 vitamin E-treated infants had birth weight of 1,500 g or less. A significant difference in incidence of neonatal sepsis (17 placebo-treated infants, 37 vitamin E-treated infants) and NEC (18 placebo-treated infants, 32 vitamin E-treated infants) was observed among infants who had been treated for eight or more days and who had developed neither sepsis nor NEC before that time. The association of vitamin E treatment with increased incidence of disease was much higher with sepsis than with NEC. The most likely reason for these observations is a pharmacologic serum vitamin E-related decrease in oxygen-dependent intracellular killing ability which results in a decreased resistance to infection in preterm infants. The data suggest that, if this occurs, it is clinically significant only in the more immature infants. In view of the known variability of absorption of oral vitamin E and the association between high serum vitamin E levels and increased incidence of sepsis and late-onset NEC reported here, it can be concluded that serum vitamin E levels must be monitored when supplemental vitamin E is administered to premature infants, especially those with birth weight 1,500 g or less. The risk-benefit ratio of long-term treatment using vitamin E at high serum levels should be clearly assessed.