Multivariate Modeling and Chemometric Resolution of Mixture Spectra in Dynamic Reaction Systems

In chemometrics, two very different classes of mathematical tools, self-modeling curve resolution and hard-modeling (first-principles) methods, have been developed to resolve pure component concentration profiles and spectra from mixture spectra recorded over time during dynamic processes in application areas from overlapped chromatographic peaks to industrial batch processes. This paper presents advantages and disadvantages of each approach via examples, and two novel applications for modeling dissolution, reaction and crystallization process in one comprehensive first-principles model. In self-modeling curve resolution (SMCR) methods, realistic constraints such as non-negativity of concentration profiles or equality constraints for known pure component spectra are imposed to produce solutions that obey the constraints and Beer’s law. In many cases, SMCR may be the only method available for resolving the pure component profiles; however, it is widely appreciated that in most circumstances, SMCR techniques do not produce unique mathematical solutions, rather a family of feasible solutions that obey boundaries imposed by the constraints. In this presentation, SMCR with a method for computing the range of feasible solutions is illustrated. An algorithm for SMCR that yields improved results by use of soft constraints with penalty functions is also described. Methods of fitting first-principles multivariate kinetic models are powerful alternatives to SMCR. Such modeling methods do not suffer from ambiguities in the resulting solutions. In process monitoring and control applications, numerical fitting of comprehensive kinetic models use dynamic information to estimate reaction rates, chemical equilibria, process states, end-points, deviations from optimal performance and can provide mechanistic information for process adjustment and optimization

Kinetic modelling of dissolution and crystallization of slurries with attenuated total reflectance UV-visible absorbance and near-infrared reflectance measurements

Slurries are often used in chemical and pharmaceutical manufacturing processes but present challenging on-line measurement and monitoring problems. In this paper, a novel multivariate kinetic modeling application is described that provides calibration-free estimates of time-resolved profiles of the solid and dissolved fractions of a substance in a model slurry system. The kinetic model of this system achieved data fusion of time-resolved spectroscopic measurements from two different kinds of fiber-optic probes. Attenuated total reflectance UV-Visible (ATR UV-Vis) and diffuse reflectance near infrared (NIR) spectra were measured simultaneously in a small-scale semi-batch reactor. A simplified comprehensive kinetic model was then fitted to the time-resolved spectroscopic data to determine the kinetics of crystallization and the kinetics of dissolution for online monitoring and quality control purposes. The parameters estimated in the model included dissolution and crystal growth rate constants, as well as the dissolution rate order. The model accurately estimated the degree of supersaturation as a function of time during conditions when crystallization took place and accurately estimated the degree of undersaturation during conditions when dissolution took place

Kinetic modeling of dissolution of salicylic acid with in situ ATR UV-vis spectroscopy

Process Analytical Technologies (PAT) recommended by the Food and Drug Administration (FDA) have significantly increased during the past years in the design, control and monitoring of pharmaceutical manufacturing processes [1]. Nowadays PAT is also commonly used in Good Manufacturing Practices (GMP). Some PAT techniques employ on-line fiber-optic sensors to acquire non-destructive measurements of physical properties, kinetic information about dissolved molecule and solid fraction of particles in slurries, in real time [2]. This study uses attenuated total reflectance ultra-violet visible (ATR UV-vis) spectroscopy for kinetic modeling of the dissolution of salicylic acid in ethanol-water within a small-scale semi-batch reactor. The dissolution model, which relies on a power-law equation, can be obtained by adding aliquots of an ethanol-water mixture into a salicylic acid slurry. Near-infrared (NIR) diffuse reflectance spectroscopy is used to detect the degree of saturation and quantify the solid fraction using a classical Kubelka-Munk transformation followed by a calibration. A temperature probe is also used to monitor heat changes involved in this dissolution. [1] Fevotte G., International Journal of Pharmaceutics 241 (2002) 263-278 [2] Gemperline et al, Analytical Chemistry 76 (2004) 2575-258

Kinetic modeling of dissolution and crystallization of batch reactions with in situ spectroscopic measurements

The use of Process Analytical Technologies (PATs) recommended by the Food and Drug Administration (FDA) has significantly increased during the past years in the design, control and monitoring of pharmaceutical or chemical manufacturing processes [1]. Nowadays PAT is also commonly used in Good Manufacturing Practices (GMPs). Some PAT techniques employ on-line fiber-optic sensors to acquire non-destructive measurements of physical properties and kinetic information of dissolved and solid fraction of molecular substances in slurries in real time [2]. The main objective of this project is to develop a global model for DuPont’s sulfonylurea coupling reaction for monitoring purposes (e.g. detect process upset, detect endpoint, and forecast changes). The global model will allow us to estimate the kinetics of the reaction, the kinetics of nucleation, the kinetics of dissolution, and the kinetics of crystallization. Before such complex model can be developed, it is necessary to conduct experiments in a simpler system (e.g. salicylic acid in water-ethanol mixture). These experiments allow us to control critical parameters and establish precisely known conditions. In addition, they are designed so the process of dissolution and the process of crystallization can be observed independently one another, and independently of chemical reactions. Consequently, we will be able to develop an appropriate model for each process in our small-scale semi-batch reactor. For the simpler system, this study uses attenuated total reflectance ultra-violet visible (ATR UV-vis) spectroscopy for kinetic modeling of the dissolution and crystallization of salicylic acid in ethanol-water. The dissolution model, which relies on a power-law equation, can be obtained by adding aliquots of an ethanol-water mixture into a salicylic acid slurry. Near-infrared (NIR) diffuse reflectance spectroscopy is used to detect the degree of saturation, quantify the solid fraction present in a slurry using a calibration, and for verification and validation purposes. A temperature probe is also used to monitor heat changes involved in this dissolution. [1] Fevotte G., International Journal of Pharmaceutics 241 (2002) 263-278 [2] Gemperline et al, Analytical Chemistry 76 (2004) 2575-258

Clinical Study of Ursodeoxycholic Acid in Barrett's Esophagus Patients

Prior research strongly implicates gastric acid and bile acids, two major components of the gastroesophageal refluxate, in the development of Barrett’s esophagus (BE) and its pathogenesis. Ursodeoxycholic acid (UDCA), a hydrophilic bile acid, has been shown to protect esophageal cells against oxidative stress induced by cytotoxic bile acids. We conducted a pilot clinical study to evaluate the clinical activity of UDCA in patients with BE. Twenty-nine BE patients received UDCA treatment at a daily dose of 13–15 mg/kg/day for six months. The clinical activity of UDCA was assessed by evaluating changes in gastric bile acid composition and markers of oxidative DNA damage (8-hydroxydeoxyguanosine, 8OHdG), cell proliferation (Ki67), and apoptosis (cleaved caspase 3, CC3) in BE epithelium. The bile acid concentrations in gastric fluid were measured by liquid chromatography-mass spectrometry. At baseline, UDCA (sum of unchanged and glycine/taurine conjugates) accounted for 18.2% of total gastric bile acids. Post UDCA intervention, UDCA increased significantly to account for 93.39% of total gastric bile acids (p<0.0001). The expression of markers of oxidative DNA damage, cell proliferation, and apoptosis was assessed in the BE biopsies by immunohistochemistry. The selected tissue biomarkers were unchanged after 6 months of UDCA intervention. We conclude that high dose UDCA supplementation for six months resulted in favorable changes in gastric bile acid composition but did not modulate selected markers of oxidative DNA damage, cell proliferation, and apoptosis in the BE epithelium

Kinetic modelling of batch slurry reactions

Slurries are widely used in important chemical and pharmaceutical manufacturing processes. In this presentation we report chemical monitoring and modeling of batch slurry reactions using optical and calorimetric measurement where key analytes are present in both the solid and liquid phases. Experimental studies were performed to: (1) Generate supersaturated solutions for the development of kinetic models of seeded crystallization, crystal growth, and dissolution; (2) Generate unseeded crystallization from homogeneous reaction mixtures for the development of kinetic models of reacting systems, including crystal growth as well as measurement of crystallization driven by cooling of reaction mixtures including the necessary corresponding solubility curves as a function of temperature; (3) Reproduce at laboratory scale, a commercially relevant slurry reaction starting with heterogeneous reaction mixture, producing a product slurry. Simultaneous UV/visible attenuated total reflectance (ATR) and NIR reflectance measurements enabled reliable characterization of reaction mixtures and detection of the onset of crystallization. Kinetic models for dissolution, seeded crystallization, unseeded crystallization, homogeneous reaction and crystallization driven by cooling were successfully developed relating the mass of substance in the solution phase and solid phase to UV/Vis ATR and NIR reflectance measurements. Reactive crystallization experiments with kinetic modeling demonstrated that simultaneous measurements of slurries with UV/Vis ATR and NIR reflectance is an effective means of characterizing the dissolved fraction and the onset of nucleation and crystallization. Work with research partner DuPont Crop Protection Science included development of a 4 L glass reactor with a recirculation loop and sampling system for off-line HPLC analysis. Several different sampling strategies were tested to develop a robust, reproducible method for off-line HPLC analysis. NIR reflectance measurements of slurry reactions at DuPont were demonstrated to be relevant to the property of interest. Kinetic models were obtained for product and reactant from NIR reflectance measurements. Reaction progression was followed by depletion of triazine heterocycle, as indicated by N-H overtone band dissipation. Spectroscopic evidence of a precipitation event was also observed and confirmed by focused beam reflectance measurements (FBRM). Kinetic models of reaction measurements provided clear resolution of two components, the solid phase reactant and solid phase product. Initial product appearance in the kinetic model concentration profile estimates corresponded temporally with the spectroscopic evidence of precipitation. Estimation of spectral profiles from kinetic models include an N-H overtone band observed around 2010nm

Understanding how education/support groups help lone mothers

<p>Abstract</p> <p>Background</p> <p>Lone-mother led families are at increased risk of psychosocial disadvantage, social isolation and mental health morbidity. Community-based programs are more accessible for families seeking assistance. We examine the experiences of eight lone mothers participating in a larger randomized controlled trial (RCT) of a community-based education/support group program using mixed methods.</p> <p>Methods</p> <p>A purposeful sample of eight mothers participating in the intervention arm of an RCT of community-based support/education groups was selected for the qualitative study. Individual interviews asked mothers about themselves and their relationships with their children before and after the group. Interviews were taped, transcribed and content analysis was used to code and interpret the data. Quantitative data collected in the RCT were used to describe these mothers.</p> <p>Results</p> <p>Mothers participating in the RCT and qualitative study experienced multiple difficulties, including financial and mood problems. These mothers reported that before participating in the group, they had shared experiences of social isolation, stigma, a sense of failure, poor relationships with their children and difficulties with financial management. After the group, mothers identified improved self-esteem, support from other mothers, improved parenting skills and improved communication with their children as outcomes of group participation.</p> <p>Conclusions</p> <p>The qualitative data revealed mothers' perceptions of specific areas that improved by participating in the group. The utility of complementary information provided by qualitative and quantitative methods in understanding program impact, as well as the need for broader assistance is noted.</p

The lipoxygenase-dependent oxygenation of lipid body membranes is promoted by a patatin-type phospholipase in cucumber cotyledons

Oilseed germination is characterized by the mobilization of storage lipids as a carbon and energy source for embryonic growth. In addition to storage lipid degradation in germinating oilseeds via the direct action of a triacylglycerol lipase (TGL) on the storage lipids, a second degradation pathway that is dependent on a specific lipid body trilinoleate 13-lipoxygenase (13-LOX) has been proposed in several plant species. The activity of this specific 13-LOX leads first to the formation of ester lipid hydroperoxides. These hydroperoxy fatty acids are then preferentially cleaved off by a TGL and serve as a substrate for glyoxysomal β-oxidation. As a prerequisite for triacylglycerol (TAG) mobilization, a partial degradation of the phospholipid monolayer and/or membrane proteins of the oil body has been discussed. Evidence has now been found for both processes: partial degradation of the proteins caleosin and oleosin was observed and simultaneously a patatin-like protein together with transient phospholipase (PLase) activity could be detected at the oil body membranes during germination. Moreover, in vitro experiments with isolated oil bodies from mature seeds revealed that the formation of 13-LOX-derived lipid peroxides in lipid body membranes is increased after incubation with the purified recombinant patatin-like protein. These experiments suggest that in vivo the degradation of storage lipids in cucumber cotyledons is promoted by the activity of a specific oil body PLase, which leads to an increased decomposition of the oil body membrane by the 13-LOX and thereby TAGs may be better accessible to LOX and TGL

Repressive Effects of Resveratrol on Androgen Receptor Transcriptional Activity

The chemopreventive effects of resveratrol (RSV) on prostate cancer have been well established; the androgen receptor (AR) plays pivotal roles in prostatic tumorigenesis. However, the exact underlying molecular mechanisms about the effects of RSV on AR have not been fully elucidated. A model system is needed to determine whether and how RSV represses AR transcriptional activity.The AR cDNA was first cloned into the retroviral vector pOZ-N and then integrated into the genome of AR-negative HeLa cells to generate the AR(+) cells. The constitutively expressed AR was characterized by monitoring hormone-stimulated nuclear translocation, DNA binding, and transcriptional activation, with the AR(-) cells serving as controls. AR(+) cells were treated with RSV, and both AR protein levels and AR transcriptional activity were measured simultaneously. Chromatin immunoprecipitation (ChIP) assays were used to detect the effects of RSV on the recruitment of AR to its cognate element (ARE).AR in the AR (+) stable cell line functions in a manner similar to that of endogenously expressed AR. Using this model system we clearly demonstrated that RSV represses AR transcriptional activity independently of any effects on AR protein levels. However, neither the hormone-mediated nucleus translocation nor the AR/ARE interaction was affected by RSV treatment.We demonstrated unambiguously that RSV regulates AR target gene expression, at least in part, by repressing AR transcriptional activity. Repressive effects of RSV on AR activity result from mechanisms other than the affects of AR nuclear translocation or DNA binding