- Quality and stability of extemporaneous pyridoxal phosphate preparations used in the treatment of paediatric epilepsy
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Objectives: To assess the pyridoxal 5′-phosphate (PLP) content and stability of extemporaneous PLP liquids prepared from dietary supplements used for the treatment of vitamin B6-dependent epilepsy. Methods: Pyridoxal 5′-phosphate liquids were prepared in accordance with the guidelines given to patients from marketed 50 mg PLP dietary capsules and tablets. The PLP content and its stability were evaluated under conditions resembling the clinical setting using reverse phase HPLC and mass spectrometry. Key findings: Pyridoxal 5′-phosphate content in most of the extemporaneously prepared liquids from dietary supplements was found to be different from the expected amount (~16–60 mg). Most of these PLP extemporaneous liquids were stable at room temperature (protected from light) after 24 h but unstable after 4 h when exposed to light. A key photodegradation product of PLP in water was confirmed as 4-pyridoxic acid 5′-phosphate (PAP). Conclusion: Pyridoxal 5′-phosphate tablets from Solgar were found to be the most reliable product for the preparation of extemporaneous PLP liquids. This work highlighted the difference between the marketed PLP dietary supplements quality and the importance of proper storage of aqueous PLP. There is a need to develop pharmaceutical forms of PLP that ensure dose accuracy and avoid potentially unsafe impurities with the aim of enhancing safety and compliance.
- Mohamed-Ahmed, Abeer H. A.,Wilson, Matthew P.,Albuera, Maedelyn,Chen, Ting,Mills, Philippa B.,Footitt, Emma J.,Clayton, Peter T.,Tuleu, Catherine
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Read Online
- SELECTIVE SOLVENT FREE PHOSPHORYLATION
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A synthetic process is provided for the preparation of phosphorylated analogs of nicotinamide riboside ("NR") having the formula (I), or salts thereof, and reduced or modified derivatives thereof, having the formula (II), wherein X-, Y1, Y2, Z1, Z2, n, R1, R2, R3, R4, R5, R6, and R7 are as defined herein. The present disclosure also relates to the preparation of phosphorylated analogs of nicotinic acid riboside ("NAR") having the formula (I), or salts thereof, and reduced or modified derivatives thereof, having the formula (II). Generally solvent-free conditions are employed using appropriate mechano-chemical techniques as described. (I) (II)
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Paragraph 0247; 0248
(2016/12/22)
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- Light-enhanced catalysis by pyridoxal phosphate-dependent aspartate aminotransferase
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The mechanisms of pyridoxal 5′-phosphate (PLP)-dependent enzymes require substrates to form covalent "external aldimine" intermediates, which absorb light strongly between 410 and 430 nm. Aspartate aminotransferase (AAT) is a prototypical PLP-dependent enzyme that catalyzes the reversible interconversion of aspartate and α-ketoglutarate with oxalacetate and glutamate. From kinetic isotope effects studies, it is known that deprotonation of the aspartate external aldimine Cα-H bond to give a carbanionic quinonoid intermediate is partially rate limiting in the thermal AAT reaction. We show that excitation of the 430-nm external aldimine absorption band increases the steady-state catalytic activity of AAT, which is attributed to the photoenhancement of Cα-H deprotonation on the basis of studies with Schiff bases in solution. Blue light (250 mW) illumination gives an observed 2.3-fold rate enhancement for WT AAT activity, a 530-fold enhancement for the inactive K258A mutant, and a 58600-fold enhancement for the PLP-Asp Schiff base in water. These different levels of enhancement correlate with the intrinsic reactivities of the Cα-H bond in the different environments, with the less reactive Schiff bases exhibiting greater enhancement. Time-resolved spectroscopy, ranging from femtoseconds to minutes, was used to investigate the nature of the photoactivation of C α-H bond cleavage in PLP-amino acid Schiff bases both in water and bound to AAT. Unlike the thermal pathway, the photoactivation pathway involves a triplet state with a Cα-H pKa that is estimated to be between 11 and 19 units lower than the ground state for the PLP-Val Schiff base in water.
- Hill, Melissa P.,Carroll, Elizabeth C.,Vang, Mai C.,Addington, Trevor A.,Toney, Michael D.,Larsen, Delmar S.
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supporting information; experimental part
p. 16953 - 16961
(2011/02/16)
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