The color purple: from royalty to laboratory, with apologies to Malachowski.

The components of the blood stain, eosin and methylene blue, were introduced by Baeyer and Caro, respectively. Methylene blue was used primarily for detecting Mycobacterium tuberculosis until Ehrlich in 1880 mixed methylene blue with acid fuchsin to produce what he termed a "neutral stain," which allowed differentiation of blood cells. Eight years later, Checin ski changed the acidic component of the dye to eosin. Plehn subsequently altered the proportions of eosin and methylene blue to produce a greater range of red and blue hues. In 1891, Malachowski and Romanowsky independently developed stains composed of eosin and "ripened" methylene blue that not only differentiated blood cells, but also demonstrated the nuclei of malarial parasites. A number of "ripening" or "polychroming" techniques were investigated by different groups, but the aqueous dye solutions produced were unstable and precipitated rapidly. Subsequently, methanol was introduced as a solvent for the dye precipitate and techniques were developed that utilized the fixative properties of the methanolic solution prior to aqueous dilution for staining. This avoided the troublesome process of heat fixation of blood films. Giemsa further improved these techniques by using more controlled methods of methylene blue demethylation. In addition, he used measured amounts of known dyes and increased dye stability by adding glycerol to the methanol solvent. With the outbreak of World War I, it became difficult to obtain German dyes outside of Germany; during the World War II, it became impossible. In their effort to improve the inferior American versions of Giemsa's stain, Lillie, Roe, and Wilcox discovered that the best staining results were obtained using pure methylene blue, one of its breakdown products (azure B) and eosin. These three substituents remain the major components of the stain to this day.

Flexibility of CuCl4-tetrahedra in bis[cinchoninium tetrachlorocuprate(II)]trihydrate single crystals. X-ray diffraction and EPR studies.

Crystal structure of bis[cinchoninium tetrachlorocuprate(II)] trihydrate, [(C19H24N2O)CuCl4]2-3H2O, has been determined by X-ray diffraction at 100 K and reexamined at 293 K. The compound crystallizes in orthorhombic system with a P2(1)2(1)2(1) space group and unit cell parameters a = 15.3031(14), b = 36.415(3), and c = 7.8341(5) A at 100 K, and Z = 4. The asymmetric unit consists of two (CuCl4)(2-) tetrahedral anions linked by hydrogen bonds to two doubly protonated cinchonine molecules and three water molecules. The tetrahedra are strongly flattened, to approximately D(2d) symmetry, with different deformation for two inequivalent (CuCl4)(2-) -ions in the asymmetric unit. The deformation of (CuCl4)(2-) and cinchoninium cations varies with temperature due to a rearrangement of the bifurcated hydrogen bond network. This is a continuous process observed as a monotonic variation of the EPR spectral parameters and the unit cell dimensions. EPR spectra show that very weak exchange coupling J(12) = 0.0030 cm(-1) operates between Cu(2+) ions within asymmetric units, corresponding to the general formula of the compound, as well as between equivalent Cu(2+) sites of different molecules, whereas the coupling is negligible between inequivalent sites. The intermolecular J(12) coupling is temperature-independent indicating that the whole asymmetric unit behaves as a magnetic unit (pseudodimer) in the whole temperature range.

Conformation of epicinchonine and cinchonine in view of their antimalarial activity: x-ray and theoretical studies.

X-ray structure analysis was carried out for a single crystal of 9-epi-10,11-dihydrocinchonine in the form of free base obtained by stereoselective interconversion of cinchonine via 9-O-tosylcinchonine. An intramolecular hydrogen bond was found between the carbinol hydroxyl group, -O12-H12, and the quinuclidine nitrogen atom, N1, with the parameters: O12...N1=2.688(3)A, O12-H12=0.84(4)A, N1...H12=2.11(4)A and O12-H12...N1=126(3) degrees. Theoretical calculations for isolated molecules of epicinchonine and cinchonine with the use of AM1 semiempirical method and comparative studies of the crystal structures have shown that the conformation of the alkaloid molecules with respect to the C8-C9 bond depends on the absolute configuration at C9. The conformation with respect to the C9-C16 bond depends on the protonation of N1 for threo but not for erythro alkaloids. It was established that the ability to form inter- or intramolecular hydrogen bonds is determined by the energetically preferred conformations of erythro and threo alkaloids, respectively. In most cases the conformations preferred for erythro alkaloids are energetically forbidden for their threo epimers and vice versa. The differences in conformation and capability to form intramolecular hydrogen bonds may explain why their antimalarial activities are incomparable.

Quinidine as a muscarinic antagonist: a structural approach.

The synthesis, spectroscopic characteristics, and single-crystal X-ray structural analysis of quitenidine methyl ester monohydrate, a derivative of the muscarinic antagonist quinidine, are presented. Quitenidine methyl ester monohydrate (C20H24N2O4.H2O) crystallizes in the orthorhombic space group P2(1)2(1)2(1), with a = 16.69(3) A, b = 12.46(2) A, c = 9.70(1) A, and Z = 4. The crystal structure was refined to a discrepancy factor (R) of 0.097. Substitution of the quinidine vinyl chain with a carboxymethyl group does not influence the conformation. The carboxymethyl group is positionally disordered, a fact that complicates refinement of the structure. The water molecule is bonded to the quinuclidine nitrogen atom, and the hydroxyl group forms an intermolecular hydrogen bond with the quinoline nitrogen atom. The molecular structure of the ester was compared with those of quinidine, quinine, and four other antimuscarinic agents. An approximately linear relationship between the distance from the nonaromatic nitrogen to the plane of the aromatic part of the molecules and the blocking potency of these agents was noted; the greater this distance, the more potent is the antagonist.

Molecular properties of Cinchona alkaloids: a theoretical approach.

In the present work, the conformation analysis, electrostatic potential calculations, and proton affinity evaluation are carried out for Cinchona alkaloids using theoretical molecular mechanics and quantum mechanical methods. The most probable conformation of the active erythro isomers at the receptor site seems to be that which enables the molecule to form intermolecular hydrogen bonds. In epiquinidine, the mutual orientation of O(12) and N(1) atoms favors intra- rather than intermolecular bonding, and this might be responsible for its inactivity. Comparison of the shape and size of the negative electrostatic potential areas provides a tentative explanation for the interaction of different erythro diastereoisomers with the same putative receptor, as well as for lack of such interaction in epiquinidine. The protonation energies calculated for cinchonidine and cinchonine confirm the higher basicity of the aliphatic N(1) as compared with that of the aromatic N(13) atom.