Managing emerging mutagenicity risks: Late stage mutagenic impurity control within the atovaquone second generation synthesis.

The mutagenic-impurity control strategy for a second generation manufacturing route to the non-mutagenic antipneumocystic agent atovaquone (2-((1R,4R)-4-(4-chlorophenyl)cyclohexyl)-3-hydroxynaphthalene-1,4-dione) 1 is described. Preliminary assessment highlighted multiple materials of concern which were largely discharged either through returning a negative bacterial mutagenicity assay or through confidence that the impurity would be purged during the downstream processing from when it was first introduced. Additional genotoxicity testing highlighted two materials of concern where initial assessment suggested that testing for these impurities at trace levels within the drug substance would be required. Following a thorough review of process purging detail, spiking and purging experimentation, and an understanding of the process parameters to which they were exposed an ICH M7 Option 4 approach could be justified for their control. The development of two 1H NMR spectroscopy methods for measurement of these impurities is also described as well as a proposed summary table for describing the underlying rationale for ICH M7 control rationales to regulators. This manuscript demonstrates that process purging of potential mutagenic impurities can be realised even when they are introduced in the later stages of a process and highlights the importance of scientific understanding rather than relying on a stage-counting approach.

Structure elucidation and spectroscopic analysis of photodegradants of the anti-rhinitis drug fluticasone furoate.

Fluticasone furoate is a novel glucocorticoid receptor agonist marketed as a treatment for seasonal and perennial allergic rhinitis. Forced degradation of fluticasone furoate under conditions of light led to a number of degradation products, the structures of which were elucidated using mass spectrometry and a range of one and two-dimensional NMR experiments. Three structures were derived, including two which involved a rearrangement of the steroid ring A to give cross-linked degradation products. The results demonstrate the applicability of a previously observed mechanism of photodegradation to fluticasone furoate.

The Vancomycin Group of Antibiotics and the Fight against Resistant Bacteria.

A last line of defence against "superbugs" are the vancomycin group antibiotics. This review describes the determination of their mode of action, and a mechanism of resistance to them. Remarkably, this mechanism of resistance can be overcome without directly modifying the binding site of the antibiotics for the cell-wall precursors of pathogenic bacteria.

Formation and identification of a degradant in chlorproguanil-dapsone-artesunate (Dacart™) tablets.

Chlorproguanil hydrochloride, dapsone and artesunate are three compounds with anti-malarial properties developed as a triple combination drug product (Dacart™) for the treatment of malarial infections. During long-term stability studies, a degradant was observed which increased with time and had the potential to limit the shelf-life of the product. Through a combination of HPLC and spectroscopic analyses, the structure of the degradant was identified to be an adduct of a fragment of artesunate with dapsone. The response factor was determined to allow an accurate assessment of its levels in drug product. The likely mechanism for its formation is postulated to be via the water-mediated degradation of artesunate to give succinic acid followed by reaction of the liberated succinic acid with dapsone. The formation of this degradant demonstrates a potential stability risk for future combination therapies incorporating artesunate. These risks are particularly pertinent to products of this type given the climatic conditions which prevail in countries where such therapies are likely to be employed.

1H chemical shifts in NMR. Part 21--prediction of the 1H chemical shifts of molecules containing the ester group: a modelling and ab initio investigation.

The 1H NMR spectra of 24 compounds containing the ester group are given and assigned. These data were used to investigate the effect of the ester group on the 1H chemical shifts in these molecules. These effects were analysed using the CHARGE model, which incorporates the electric field, magnetic anisotropy and steric effects of the functional group for long-range protons together with functions for the calculation of the two- and three-bond effects. The effect of the ester electric field was given by considering the partial atomic charges on the three atoms of the ester group. The anisotropy of the carbonyl group was reproduced with an asymmetric magnetic anisotropy acting at the midpoint of the carbonyl bond with values of Deltachi(parl) and Deltachi(perp) of 10.1 x 10(-30) and -17.1 x 10(-30) cm3 molecule(-1). An aromatic ring current (=0.3 times the benzene ring current) was found to be necessary for pyrone but none for maleic anhydride. This result was confirmed by GIAO calculations. The observed 1H chemical shifts in the above compounds were compared with those calculated by CHARGE and the ab initio GIAO method (B3LYP/6-31G**). For the 24 compounds investigated with 150 1H chemical shifts spanning a range of ca 10 ppm, the CHARGE model gave an excellent r.m.s. error (obs - calc) of <0.1 ppm. The GIAO calculations gave a very reasonable r.m.s. error of ca 0.2 ppm although larger deviations of ca 0.5 ppm were observed for protons near to the electronegative atoms. The accurate predictions of the 1H chemical shifts given by the CHARGE model were used in the conformational analysis of the vinyl esters methyl acrylate and methyl crotonate. An illustration of the use of the CHARGE model in the prediction of the 1H spectrum of a complex organic molecule (benzochromen-6-one) is also given.

HPLC-SPE-NMR in pharmaceutical development: capabilities and applications.

High-performance liquid chromatography-solid phase extraction-NMR spectroscopy (HPLC-SPE-NMR) has recently become commercially available and has been evaluated with regard to its applicability in a pharmaceutical environment. The addition of an automated SPE unit to an HPLC-NMR system for peak trapping results in an improved NMR signal-to-noise ratio (S/N) and also has other practical advantages. The trapping efficiency is shown to depend on compound polarity and is highest for compounds eluting late on reversed-phase HPLC systems. Multiple peak trapping further increases the S/N, again with the best results for less polar compounds. For polar compounds, multiple peak trapping resulted in no S/N gain as the amount of material retained on the SPE cartridge was equivalent to that from a single injection. When compared with conventional HPLC-NMR, a S/N gain of up to five-fold could be achieved for some compounds in a single trapping step. A major advantage of the technique is the independence of the chromatographic step from the NMR step, resulting in greater versatility than conventional HPLC-NMR in the HPLC solvents and NMR solvents that can be used. Practical applications from both drug metabolite and drug impurity identification are presented.

Noncovalent interactions: defining cooperativity. Ligand binding aided by reduced dynamic behavior of receptors. Binding of bacterial cell wall analogues to ristocetin A.

Changes in the relative populations of the monomer and asymmetric dimer forms of ristocetin A, upon binding of two molecules of ligand, suggest that ligand binding is negatively cooperative with respect to dimerization. However, strong hydrogen bonds formed in the binding sites of the ligands are reinforced in the dimer relative to the monomer, and the barrier to dissociation of the dimer is increased upon binding of the ligands. It is concluded that the interactions which are common in the binding of both ligands are made with positive cooperativity with respect to those involved in dimerization. The conclusions are relevant to the binding of ligands to proteins, where ligand binding energy can be derived from stabilization of the protein in its ligand-bound form.