A New Method for Producing Pharmaceutical Co-crystals: Laser Irradiation of Powder Blends

In this work, a high-power CO2 laser was used to irradiate powder blends of co-crystal formers, with the specific aim of trying to cause recrystallization to a co-crystal structure. By varying the power and raster speed of the laser, it was found that sufficient thermal energy could be imparted to the powder to cause molecular rearrangement. It was possible to form co-crystals of caffeine with oxalic acid and caffeine with malonic acid. Interestingly, it was found that, to form co-crystals successfully, the coformers needed to sublime to an appreciable extent, which indicates that the mechanism of rearrangement involves interaction and nucleation in the vapor phase. Laser irradiation thus offers a new route to creation of pharmaceutical co-crystals and a potentially rapid screen for likely co-crystal formation between coforming pairs

Laser irradiation to produce amorphous pharmaceuticals

Using a high-power CO2 laser to irradiate powder beds, it was possible to induce phase transformation to the amorphous state. Irradiation of a model drug, indometacin, resulted in formation of a glass. Varying the settings of the laser (power and raster speed) was shown to change the physicochemical properties of the glasses produced and all irradiated glasses were found to be more stable than a reference glass produced by melt-quenching. Irradiation of a powder blend of paracetamol and polyvinylpyrrolidone K30 was found to produce a solid amorphous dispersion. The results suggest that laser-irradiation might be a useful method for making amorphous pharmaceuticals

Effect of CO2 laser irradiation on Eudragit® L100-55, L100, and S100 coatings to modify drug release

The aim of this work was to investigate the use of carbon dioxide (CO2 ) laser irradiation to modify three types of pH-dependent Eudragit® (L100-55, L100, and S100) enteric coats with the aim of modulating drug release kinetics from the tablet cores. CO2 laser irradiation causes rapid melting and resolidification/vaporization of materials locally and precisely through the absorption of infrared energy and so can potentially disrupt the barrier integrity and function of enteric coats. It was successfully utilized to shorten the lag time of drug release (T50% and T80% ) during dissolution testing. These changes were mainly caused either by pore formation on the surface of the coating and/or loosening of the film coat. In addition, changes in mechanical properties (Young’s modulus and tensile strengths) and shifted IR peaks of the irradiated coatings were found, which correlated with drug release rates. This work is a proof-of-concept of tailoring drug release profiles by adjusting the power of the laser energy which could be useful for the modification of drug release for personalized medicines

phot1 inhibition of ABCB19 primes lateral auxin fluxes in the shoot apex required for phototropism

It is well accepted that lateral redistribution of the phytohormone auxin underlies the bending of plant organs towards light. In monocots, photoreception occurs at the shoot tip above the region of differential growth. Despite more than a century of research, it is still unresolved how light regulates auxin distribution and where this occurs in dicots. Here, we establish a system in Arabidopsis thaliana to study hypocotyl phototropism in the absence of developmental events associated with seedling photomorphogenesis. We show that auxin redistribution to the epidermal sites of action occurs at and above the hypocotyl apex, not at the elongation zone. Within this region, we identify the auxin efflux transporter ATP-BINDING CASSETTE B19 (ABCB19) as a substrate target for the photoreceptor kinase PHOTOTROPIN 1 (phot1). Heterologous expression and physiological analyses indicate that phosphorylation of ABCB19 by phot1 inhibits its efflux activity, thereby increasing auxin levels in and above the hypocotyl apex to halt vertical growth and prime lateral fluxes that are subsequently channeled to the elongation zone by PIN-FORMED 3 (PIN3). Together, these results provide new insights into the roles of ABCB19 and PIN3 in establishing phototropic curvatures and demonstrate that the proximity of light perception and differential phototropic growth is conserved in angiosperm

Cellular efflux of auxin catalyzed by the Arabidopsis MDR/PGP transporter AtPGP1

Directional transport of the phytohormone auxin is required for the establishment and maintenance of plant polarity, but the underlying molecular mechanisms have not been fully elucidated. Plant homologs of human multiple drug resistance/P-glycoproteins (MDR/PGPs) have been implicated in auxin transport, as defects in MDR1 (AtPGP19) and AtPGP1 result in reductions of growth and auxin transport in Arabidopsis (atpgp1, atpgp19), maize (brachytic2) and sorghum (dwarf3). Here we examine the localization, activity, substrate specificity and inhibitor sensitivity of AtPGP1. AtPGP1 exhibits non-polar plasma membrane localization at the shoot and root apices, as well as polar localization above the root apex. Protoplasts from Arabidopsis pgp1 leaf mesophyll cells exhibit reduced efflux of natural and synthetic auxins with reduced sensitivity to auxin efflux inhibitors. Expression of AtPGP1 in yeast and in the standard mammalian expression system used to analyze human MDR-type proteins results in enhanced efflux of indole-3-acetic acid (IAA) and the synthetic auxin 1-naphthalene acetic acid (1-NAA), but not the inactive auxin 2-NAA. AtPGP1-mediated efflux is sensitive to auxin efflux and ABC transporter inhibitors. As is seen in planta, AtPGP1 also appears to mediate some efflux of IAA oxidative breakdown products associated with apical sites of high auxin accumulation. However, unlike what is seen in planta, some additional transport of the benzoic acid is observed in yeast and mammalian cells expressing AtPGP1, suggesting that other factors present in plant tissues confer enhanced auxin specificity to PGP-mediated transport. © 2005 Blackwell Publishing Ltd

Brachytic2/ZmABCB1 functions in IAA export from intercalary meristems

Dwarfism traits in Zea mays are regulated by multiple factors including the hormone auxin. Dwarf brachytic2 (br2) mutants harbour lesions in the gene encoding an orthologue of Arabidopsis thaliana ABCB1 which functions in auxin efflux out of meristematic regions in the shoot and root. br2 mesocotyls and coleoptiles exhibit reduced auxin transport. However, the dwarf stature of br2 derives from shortened lower internodes whilst the upper portion of the plant is completely normal. As such, it is counter-intuitive to attribute br2 dwarfism exclusively to reduced auxin export out of the shoot apex. Arabidopsis abcb1 mutants exhibit only minor reductions in auxin transport and plant height unless combined with mutations in the ABCB19 auxin transporter. Phylogenetic modelling analysis excludes the possibility that BR2 is more closely related to ABCB19 which has three more closely related orthologues in maize. BR2 is expressed in nodal meristems, and analyses of auxin transport and content indicate that BR2 function in these grass-specific tissues is analogous to ABCB1 function in the shoot and root apex of Arabidopsis. These results indicate that ABCB1/BR2 function is conserved between dicots and monocots, but also suggests that this function must be understood in the context of the segmental organization of grass plants

Auxin influx inhibitors 1-NOA, 2-NOA, and CHPAA interfere with membrane dynamics in tobacco cells

The phytohormone auxin is transported through the plant body either via vascular pathways or from cell to cell by specialized polar transport machinery. This machinery consists of a balanced system of passive diffusion combined with the activities of auxin influx and efflux carriers. Synthetic auxins that differ in the mechanisms of their transport across the plasma membrane together with polar auxin transport inhibitors have been used in many studies on particular auxin carriers and their role in plant development. However, the exact mechanism of action of auxin efflux and influx inhibitors has not been fully elucidated. In this report, the mechanism of action of the auxin influx inhibitors (1-naphthoxyacetic acid (1-NOA), 2-naphthoxyacetic acid (2-NOA), and 3-chloro-4-hydroxyphenylacetic acid (CHPAA)) is examined by direct measurements of auxin accumulation, cellular phenotypic analysis, as well as by localization studies of Arabidopsis thaliana L. auxin carriers heterologously expressed in Nicotiana tabacum L., cv. Bright Yellow cell suspensions. The mode of action of 1-NOA, 2-NOA, and CHPAA has been shown to be linked with the dynamics of the plasma membrane. The most potent inhibitor, 1-NOA, blocked the activities of both auxin influx and efflux carriers, whereas 2-NOA and CHPAA at the same concentration preferentially inhibited auxin influx. The results suggest that these, previously unknown, activities of putative auxin influx inhibitors regulate overall auxin transport across the plasma membrane depending on the dynamics of particular membrane vesicles

Cytokinin acts through the auxin influx carrier AUX1 to regulate cell elongation in the root

Hormonal interactions are critical for plant development. In Arabidopsis, cytokinins inhibit root growth through effects on cell proliferation and cell elongation. Here we define key mechanistic elements in a regulatory network by which cytokinin inhibits root cell elongation in concert with the hormones auxin and ethylene. The auxin importer AUX1 functions as a positive regulator of cytokinin responses in the root, AUX1 mutants specifically affecting the ability of cytokinin to inhibit cell elongation but not cell proliferation. AUX1 is required for cytokinin-dependent changes of auxin activity in the lateral root cap associated with the control of cell elongation. Cytokinin regulates root cell elongation through ethylene-dependent and independent mechanisms, both hormonal signals converging on AUX1 as a regulatory hub. An autoregulatory circuit is identified involving the control of ARR10 and AUX1 expression by cytokinin and auxin, this circuit potentially functioning as an oscillator to integrate the effects of these two hormones. Taken together, our results uncover several regulatory circuits controlling interactions of cytokinin with auxin and ethylene, and support a model in which cytokinin regulates shootward auxin transport to control cell elongation and root growth