Selection of an analytical method for evaluating bovine serum albumin concentrations in pharmaceutical polymeric formulations

Bovine serum albumin (BSA) is a commonly used model protein in the development of pharmaceutical formulations. In order to assay its release from various dosage forms, either the bicinchoninic acid (BCA) assay or a more specific size-exclusion high performance liquid chromatography (SE-HPLC) method are commonly employed. However, these can give erroneous results in the presence of some commonly-used pharmaceutical excipients. We therefore investigated the ability of these methods to accurately determine BSA concentrations in pharmaceutical formulations that also contained various polymers and compared them with a new and compared with a new reverse-phase (RP)–HPLC technique. We found that the RP-HPLC technique was the most suitable method. It gave a linear response in the range of 0.5 -100 µg/ml with a correlation coefficient of 0.9999, a limit of detection of 0.11 µg/ml and quantification of 0.33 µg/ml. The performed ‘t’ test for the estimated and theoretical concentration indicated no significant difference between them providing the accuracy. Low % relative standard deviation values (0.8-1.39%) indicate the precision of the method. Furthermore, the method was used to quantify in vitro BSA release from polymeric freeze-dried formulations.

Langerhans cell (LC) proliferation mediates neonatal development, homeostasis, and inflammation-associated expansion of the epidermal LC network.

Most tissues develop from stem cells and precursors that undergo differentiation as their proliferative potential decreases. Mature differentiated cells rarely proliferate and are replaced at the end of their life by new cells derived from precursors. Langerhans cells (LCs) of the epidermis, although of myeloid origin, were shown to renew in tissues independently from the bone marrow, suggesting the existence of a dermal or epidermal progenitor. We investigated the mechanisms involved in LC development and homeostasis. We observed that a single wave of LC precursors was recruited in the epidermis of mice around embryonic day 18 and acquired a dendritic morphology, major histocompatibility complex II, CD11c, and langerin expression immediately after birth. Langerin+ cells then undergo a massive burst of proliferation between postnatal day 2 (P2) and P7, expanding their numbers by 10–20-fold. After the first week of life, we observed low-level proliferation of langerin+ cells within the epidermis. However, in a mouse model of atopic dermatitis (AD), a keratinocyte signal triggered increased epidermal LC proliferation. Similar findings were observed in epidermis from human patients with AD. Therefore, proliferation of differentiated resident cells represents an alternative pathway for development in the newborn, homeostasis, and expansion in adults of selected myeloid cell populations such as LCs. This mechanism may be relevant in locations where leukocyte trafficking is limited.

Reduction of severe bovine serum associated matrix effects on carboxymethylated dextran coated biosensor surfaces

Surface plasmon resonance (SPR) based biosensor technology has been widely used in life science research for many applications. While the advantages of speed, ruggedness, versatility, sensitivity and reproducibility are often quoted, many researchers have experienced severe problem of non-specific binding (NSB) to chip surfaces when performing analysis of biological samples Such as bovine serum. Using the direct measurement of the bovine protein leptin, present in bovine serum samples as a model, a unique buffering system has been developed and optimised which was able to significantly reduce the non-specific interactions of bovine serum components with the carboxymethyl dextran chip (CM5) surface on a Biacore SPR The developed NSB buffering system comprised of HBS-EP buffer, containing 0.5 M NaCl, 0.005% CM-dextran pH 9.0. An average NSB reduction (n = 20) of 85.9% and 87.3% was found on an unmodified CM5 surface and a CM5 with bovine leptin immobilised on the chip surface, respectively. A reduction in NSB of up to 94% was observed on both surfaces. The concentration of the constitutive components and pH of the buffer were crucial in achieving this outcome. (C) 2008 Elsevier B.V. All rights reserved.

Detection of glucocorticoid bioactivity in bovine urine samples using a reporter gene assay.

The illegal use of anabolic substances in the meat producing industry is an ongoing problem due to the continual production of new synthetic compounds and/or the practice of low-level cocktail administration to avoid detection by the surveillance schemes of EU member states National Plan surveillance systems.

We present a highly sensitive reporter gene assay and sample extraction procedure based on a two step solid phase extraction and high performance liquid chromatography, developed for the detection of glucocorticoid abuse in bovine urine. The assay is capable of detecting compounds with glucocorticoid bioactivity and is extremely sensitive with an EC50 of 0.79 ng mL-1 for dexamethasone. New or unknown compounds with glucocorticoid bioactivity and low-level cocktail mixtures are detectable by this assay.

Cross-reactivity data for a range of 11ß-hydroxyglucocorticoids has been provided. This assay shows low interference from the 11-keto prohormones and other steroidal hormones. The assay may be suitable for application in other matrices such as hair. In conclusion this screening assay offers advantages over existing analytical techniques.

Quasiexperimental Study of the Effects of Antibiotic Use, Gastric Acid-Suppressive Agents, and Infection Control Practices on the Incidence of Clostridium difficile-Associated Diarrhea in Hospitalized Patients

The objective of this study was to evaluate the effects of antimicrobial drug use, gastric acid-suppressive agent use, and infection control practices on the incidence of Clostridium difficile-associated diarrhea (CDAD) in a 426-bed general teaching hospital in Northern Ireland. The study was retrospective and ecological in design. A multivariate autoregressive integrated moving average (time-series analysis) model was built to relate CDAD incidence with antibiotic use, gastric acid-suppressive agent use, and infection control practices within the hospital over a 5-year period (February 2002 to March 2007). The findings of this study showed that temporal variation in CDAD incidence followed temporal variations in expanded-spectrum cephalosporin use (average delay = 2 months; variation of CDAD incidence = 0.01/100 bed-days), broad-spectrum cephalosporin use (average delay = 2 months; variation of CDAD incidence = 0.02/100 bed-days), fluoroquinolone use (average delay = 3 months; variation of CDAD incidence = 0.004/100 bed-days), amoxicillin-clavulanic acid use (average delay = 1 month; variation of CDAD incidence = 0.002/100 bed-days), and macrolide use (average delay = 5 months; variation of CDAD incidence = 0.002/100 bed-days). Temporal relationships were also observed between CDAD incidence and use of histamine-2 receptor antagonists (H2RAs; average delay = 1 month; variation of CDAD incidence = 0.001/100 bed-days). The model explained 78% of the variance in the monthly incidence of CDAD. The findings of this study highlight a temporal relationship between certain classes of antibiotics, H2RAs, and CDAD incidence. The results of this research can help hospitals to set priorities for restricting the use of specific antibiotic classes, based on the size-effect of each class and the delay necessary to observe an effect.

Neonatal control of sucking pressures

Human newborns appear to regulate sucking pressure when bottle feeding by employing, with similar precision, the same principle of control evidenced by adults in skilled behavior, such as reaching (Lee et al., 1998a). In particular, the present study of 12 full-term newborn infants indicated that the intraoral sucking pressures followed an internal dynamic prototype - an intrinsic tau-guide. The intrinsic tau-guide, a recent hypothesis of general tau theory is a time-varying quantity, tau(g), assumed to be generated within the nervous system. It corresponds to some quantity (e.g., electrical charge), chang ing with a constant second-order temporal derivative from a rest level to a goal level, in the sense that tau(g) equals tau of the gap between the quantity and its goal level at each time t. (tau of a gap is the rime-to-closure of the gap at the current closure-rate.) According to the hypoth esis, the infant senses tau(p), the tau of the gap between the current intraoral pressure and its goal level, and regulates intraoral pressure so that tau(p) and tau(g) remain coupled in a constant ratio, k; i.e., tau(p) = k tau(g). With k in the range 0-1, the tau-coupling would result in a bell-shaped rate of change pressure profile, as was, in fact, found. More specifically, the high mean r(2) values obtained when regressing tau(p) on tau(g), for both the increasing and decreasing suction periods of the infants' suck, supported a strong tau-coupling between tau(p) and tau(g). The mean k values were significantly higher In the Increasing suction period, indicating that the ending of the movement was more forceful, a finding which makes sense given the different functions of the two periods of the suck.