Optimization and validation of high-performance chromatographic condition for simultaneous determination of adapalene and benzoyl peroxide by response surface methodology.

The aim of this study was to develop a simple and reliable high-performance chromatographic (HPLC) method for simultaneous analysis of adapalene and benzoyl peroxide in pharmaceutical formulation by response surface methodology (RSM). An optimized mobile phase composed of acetonitrile, tetrahydrofuran and water containing 0.1% acetic acid at a ratio of 25:50:25 by volume was successfully predicted by using RSM. An isocratic separation was achieved by using the condition. Furthermore, the analytical method was validated in terms of specificity, linearity, accuracy and precision in a range of 80% to 120% of the expected concentration. Finally, the method was successfully applied to the analysis of a commercial product.

Formulation optimization of arecoline patches.

The response surface methodology (RSM) including polynomial equations has been used to design an optimal patch formulation with appropriate adhesion and flux. The patch formulations were composed of different polymers, including Eudragit RS 100 (ERS), Eudragit RL 100 (ERL) and polyvinylpyrrolidone K30 (PVP), plasticizers (PEG 400), and drug. In addition, using terpenes as enhancers could increase the flux of the drug. Menthol showed the highest enhancement effect on the flux of arecoline.

Isotretinoin oil-based capsule formulation optimization.

The purpose of this study was to develop and optimize an isotretinoin oil-based capsule with specific dissolution pattern. A three-factor-constrained mixture design was used to prepare the systemic model formulations. The independent factors were the components of oil-based capsule including beeswax (X1), hydrogenated coconut oil (X2), and soybean oil (X3). The drug release percentages at 10, 30, 60, and 90 min were selected as responses. The effect of formulation factors including that on responses was inspected by using response surface methodology (RSM). Multiple-response optimization was performed to search for the appropriate formulation with specific release pattern. It was found that the interaction effect of these formulation factors (X1X2, X1X3, and X2X3) showed more potential influence than that of the main factors (X1, X2, and X3). An optimal predicted formulation with Y(10 min), Y(30 min), Y(60 min), and Y(90 min) release values of 12.3%, 36.7%, 73.6%, and 92.7% at X1, X2, and X3 of 5.75, 15.37, and 78.88, respectively, was developed. The new formulation was prepared and performed by the dissolution test. The similarity factor f2 was 54.8, indicating that the dissolution pattern of the new optimized formulation showed equivalence to the predicted profile.

Thermosensitive liposomes entrapping iron oxide nanoparticles for controllable drug release.

Iron oxide nanoparticles can serve as a heating source upon alternative magnetic field (AMF) exposure. Iron oxide nanoparticles can be mixed with thermosensitive nanovehicles for hyperthermia-induced drug release, yet such a design and mechanism may not be suitable for controllable drug release applications in which the tissues are susceptible to environmental temperature change such as brain tissue. In the present study, iron oxide nanoparticles were entrapped inside of thermosensitive liposomes for AMF-induced drug release while the environmental temperature was maintained at a constant level. Carboxyfluorescein was co-entrapped with the iron oxide nanoparticles in the liposomes as a model compound for monitoring drug release and environmental temperature was maintained with a water circulator jacket. These experiments have been successfully performed in solution, in phantom and in anesthetized animals. Furthermore, the thermosensitive liposomes were administered into rat forearm skeletal muscle, and the release of carboxylfluorescein triggered by the external alternative magnetic field was monitored by an implanted microdialysis perfusion probe with an on-line laser-induced fluorescence detector. In the future such a device could be applied to simultaneous magnetic resonance imaging and non-invasive drug release in temperature-sensitive applications.

Experimental spinal cord injury induced an increase of extracellular ascorbic acid concentration in anesthetized rats: a microdialysis study.

Ascorbic acid plays important roles in mammalian central nervous system. We employed an on-line analytical system to monitor the extracellular ascorbic acid concentrations in anesthetized rat spinal cord before and after the experimental injury. A microdialysis probe (216 microm od, 200 microm id, 3 mm in length) was implanted into an anesthetized rat spinal cord (Thoratic-12). Microdialysis perfusate (2 microl/min) was collected in the sample loop (20 microl) of an on-line injector for direct injection onto a High Performance Liquid Chromatography (HPLC) system equipped with an electrochemical detector. Normal ascorbic acid concentrations in the spinal cord extracellular fluids ranged from 1.8 microM to 10.8 microM (mean +/- S.D. 5.6 +/- 2.4 microM, n = 8). The experimental spinal cord injury, induced by a lesion at T-10, gradually yet significantly increased the extracellular ascorbic acid levels. The effect of exogenous glutamate perfusion (0.2 mM, 2 mM, and 20 mM) through the microdialysis probe also increased the extracellular ascorbic acid concentrations in a dose dependent manner. These results suggested that the injury-induced ascorbic acid accumulation may result from elevated extracellular glutamate levels that are commonly observed in spinal cord injury. This on-line, continuous and automatic monitoring system can be applied to future investigations on the roles of ascorbic acid in spinal cord injuries.

In vivo monitoring of fluorescent nanosphere delivery in anesthetized rats using an implantable fiber-optic microprobe.

An implantable needle-type fiber-optic microprobe was constructed to monitor in vivo fluorescent substances in anesthetized rats. This fiber-optic microprobe was composed of coaxial optical fibers that were catheterized using a thin-wall tube of stainless steel (o.d. approximately 400 microm; i.d. approximately 300 microm). When the fiber-optic microprobe was placed in solutions containing various concentrations of fluorescent nanospheres (20 nm), either in the presence or in the absence of 10% Lipofundin acting as an optical phantom, we observed nanosphere concentration-dependent responses of the fluorescence intensity. The microprobe was then implanted into the livers and brains of anesthetized rats to monitor the in situ extravasation of preadministered fluorescent nanospheres from vasculature following the hepatic and cerebral ischemic insults. Both types of ischemic insults showed immediate increases in fluorescent intensities when 20-nm fluorescent nanosphere were administered, but neither ischemic insult induces such an increase when we administered 1000-nm fluorescent nanospheres. Additional experiments can be performed to further narrow the size range of the increase in blood vessel permeability following ischemic insult; such "size" information may be valuable when formulating drugs for optimal local delivery. Although a wide variety of fluorescent substances are used intensively for in vitro biological studies, the in vivo and in situ monitoring of these substances is studied much less often, probably because of difficulties in the efficient assembly of miniaturized fiber optics to detect the relatively weak fluorescence signal arising within such a turbid medium as tissue. To our knowledge, the use of our implantable fiber-optic microprobe is the first minimally invasive technique capable of investigating the "size window" of vascular permeability for the in vivo delivery of nanospheres in both ischemic livers and brains.

Nanoparticle-based in vivo investigation on blood-brain barrier permeability following ischemia and reperfusion.

The blood-brain barrier (BBB) represents a significant impediment to a large variety of central nervous system-active agents. In the current study, we applied fluorescent polystyrene nanospheres (20 nm) to study the BBB permeability following cerebral ischemia and reperfusion. A microdialysis probe was implanted in the cerebral cortex of an anesthetized rat injected with fluorescent polystyrene nanospheres. The circulating nanospheres extravasating to the brain extracellular fluids were collected by the probe. Fluorescence intensity in the microdialysates throughout the course of cerebral ischemia/reperfusion was measured. Cerebral ischemia and reperfusion induced transient accumulations of extracellular nanospheres in the brain. The accumulation of nanospheres may result from their extravasation from the blood vessels. The concurrent cerebral oxygen levels monitored using oxygen-dependent quenching of phosphorescence decreased following ischemia and returned to their original levels after reperfusion. In conclusion, we demonstrated that high temporal resolution measurements of BBB permeability in vivo can be obtained using fluorescence polystyrene nanospheres and that these data correlate with changes of cerebral oxygen concentration. This present investigation indicates that nanoparticles have potential clinical applications involving drug delivery and determination of therapeutic efficacy and on-site diagnosis.

Ionotropic glutamate receptors are involved in malondialdehyde production in anesthetized rat brain cortex: a microdialysis study.

Elevated extracellular glutamate levels can increase malondialdehyde production in the brains of anesthetized rats. Thus, we investigated whether ionotropic glutamate receptors are involved in glutamate-induced malondialdehyde production. A microdialysis probe was implanted in the brain cortex of anesthetized rats. The malondialdehyde level in microdialysates was analyzed using an HPLC system. Three different ionotropic glutamate receptor agonists were used. At a concentration of 1.5 mM alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid hydrobromide (AMPA, a selective AMPA receptor agonist) induced a dramatic increase in extracellular malondialdehyde production (as much as 14-fold relative to the basal value). N-Methyl-D-aspartic acid (NMDA, a selective NMDA receptor agonist) also induced an increase in extracellular malondialdehyde production; however, the increase was not as much as that observed in the perfusion of AMPA receptor agonist. Kainic acid (a selective kainate receptor agonist) did not significantly increase malondialdehyde production. When co-perfused with L-trans-pyrrolidine-2,4-dicarboxylate (PDC; 31.4 mM), a glutamate uptake transport inhibitor that can increase the extracellular glutamate levels, AMPA receptor antagonist [1-(4-aminophenyl)4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine hydrochloride, 1.0 mM] can significantly reduce PDC-induced malondialdehyde production. Although NMDA receptor antagonist [(5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate, MK801] also can decrease the PDC-induced malondialdehyde production, it was not as effective as the AMPA receptor antagonist. These results suggest that ionotropic receptors are involved in the glutamate-induced increase in malondialdehdye production. Specifically, AMPA receptor seems to be predominant in the glutamate-induced malondialdehdye production in anesthetized rat brain cortex.