Sequential Amplification of Amino Acid Enantiomeric Excess by Conglomerate and Racemic Compound: Plausible Prebiotic Route Towards Homochirality.

The strong link observed between the detection of unstable or metastable racemic compounds in the systems studied and the maximum entrainment effect attainable during preferential crystallization (PC) (assessed by the maximum value of the enantiomeric excess of the mother solution at the end of PC (eefmax)) is confirmed by the results obtained with the simultaneous resolution of enantiomorphous p and p′ salts of (±)-ephedrine and (±)-mandelic acid (eefmax = 10.6% with no racemic compound detected). The detection of such a racemic compound indicates that strong heterochiral interactions at one or more (hkl) crystal–mother liquor interfaces are favorable to the 2D nucleation of the counter enantiomer as soon as its local supersaturation at these interfaces (β*hkl) attains a threshold above which the desorption of a docked counter-enantiomer molecule is no longer probable. Therefore, the mode and the rate of stirring must be optimized in order to keep β*hkl as close as possible to the overall supersaturation of the counter enantiomer β*bulk, while minimizing the damaging effects on the particles. The crystal structures of an unstable racemic compound and of a new polymorphic form of 1-phenylethylammonium hydratropate salts are reported and contribute to the understanding of the limited entrainment effect (eefmax = 5.2%).

New studies on the reciprocal quaternary system (±)-2-phenylpropionic acid [(±)-1]–(±)-α-methylbenzylamine [(±)-2]–ethanol confirm the presence of a stable conglomerate (p and p′ salts), but polymorphism of the n salt as well as an unstable racemic compound have been detected. The kinetic parameters of the irreversible transformation of the unstable racemic compound into the stable conglomerate have been determined from experimental X-ray powder diffraction data. The simultaneous resolution of (±)-1 and (±)-2 by means of preferential crystallisation (auto-seeded process) of the stable pair of enantiomorphous salts, was achieved. However, the entrainment effect (given by the maximum enantiomeric excess of the counter enantiomer in the mother solution, reached at the end of the stereoselective crystallisation (eefmax = 5.2%)) is limited. This is consistent with the existence of the unstable racemic compound, as accounted for in a recent model of molecular interactions occurring at the crystal–mother solution interface in the course of preferential crystallisation. An extended version of this model rationalises a kinetic advantage of the crystal growth rate of racemic compounds over the conglomerates as well as a large supersaturation capacity of the mother solution in such a reciprocal quaternary system (±)-acid, (±)-base and solvent.

The application of preferential crystallization is at present limited to conglomerate forming systems, which cover only a minor part of chiral substances. In this paper, a hybrid process is proposed that extends the applicability of the preferential crystallization principle to the more common racemic compound forming systems. It comprises a preliminary (e.g., chromatographic) enantiomeric enrichment step and preferential crystallization to finally produce the desired pure enantiomer(s). The applicability of preferential crystallization to racemic compounds is demonstrated on the example of mandelic acid as a model system. Direct monitoring of the separation progress is performed using combined online polarimetry and online density measurements. A cyclic crystallization process, which provides alternating the pure mandelic acid enantiomer and the racemic compound, is feasible and allows the resolution of rac-mandelic acid as part of the proposed hybrid approach.

The relationship of racemic structure to resolution by preferential crystallization was discussed on the ternary phase diagrams of systems containing optical isomers and a solvent. From these discussions, it was concluded that the resolution by this method is applicable only to racemic mixtures, and that racemic compounds and also racemic solid solutions can not be resolved. In addition, we reexamined the resolution of 3, 3-diethyl-5-methyl-2, 4-dioxopiperidine reported to be resolved by this method in spite of the formation of a racemic solid solution, and found it impossible to resolve this compound by preferential crystallization.

Resolution of 5-hydroxymethyl-2-oxazolidinone by preferential crystallization and investigations on the nature of the racemates of some 2-oxazolidinone derivatives

Recently the feasibility of preferential crystallization for enantioseparation of racemic compound forming systems has been demonstrated (Lorenz et al., Application of preferential crystallization to resolve racemic compounds in a hybrid process. Chirality 2006;18:828-840; Polenske et al., Separation of the propranolol hydrochloride enantiomers by preferential crystallization: thermodynamic basis and experimental verification. Cryst Growth Des 2007;7:1628-1634). Here, the development and the potential of an efficient separation process operated via two different techniques of preferential crystallization are studied: (1) seeded isothermal preferential crystallization and (2) auto-seeded polythermal preferential crystallization. Both techniques were investigated in the batch and in the cyclic operation mode. On the example of mandelic acid as a typical racemic compound forming system, it is demonstrated that a cyclic auto-seeded polythermal process is feasible and significantly more efficient than the seeded isothermal one.

Additive-assisted preferential crystallization of racemic component: A case of norvaline

We tap into an unexplored area of preferential crystallization, being the first to develop simultaneous chiral resolution of two racemic compounds by preferential cocrystallization. We highlight how the two racemic compounds RS-mandelic acid (MAN) and RS-etiracetam (ETI) can be combined together as enantiospecific R-MAN⋅R-ETI and S-MAN⋅S-ETI cocrystals forming a stable conglomerate system and subsequently develop a cyclic preferential crystallization allowing to simultaneous resolve both compounds. The developed process leads to excellent enantiopurity both for etiracetam (ee>98 %) and mandelic acid (ee≈95 %) enantiomers.

We tap into an unexplored area of preferential crystallization, being the first to develop simultaneous chiral resolution of two racemic compounds by preferential cocrystallization. We highlight how the two racemic compounds RS‐mandelic acid (MAN) and RS‐etiracetam (ETI) can be combined together as enantiospecific R‐MAN⋅R‐ETI and S‐MAN⋅S‐ETI cocrystals forming a stable conglomerate system and subsequently develop a cyclic preferential crystallization allowing to simultaneous resolve both compounds. The developed process leads to excellent enantiopurity both for etiracetam (ee>98 %) and mandelic acid (ee≈95 %) enantiomers.

Aspartic acid is an amino acid present in the modern proteins, however, is considered a primitive amino acid hence its importance in prebiotic chemistry experiments studies. In some works of prebiotic chemistry have been studied the synthesis and the stability of organic matter under high energy sources, and the role of clays has been highlighted due to clays that can affect the reaction mechanisms in the radiolytic processes. The present work is focused on the study of the role of Namontmorillonite in the gamma radiolysis processes of L-aspartic acid. Gamma radiolysis processes were carried out in three different systems a) L-aspartic acid in aqueous solution; b) L-aspartic acid in solid-state; and c) L-aspartic acid adsorbed into Na-montmorillonite. L-aspartic acid was analyzed by high-performance liquid chromatography−electrospray ionization−mass spectrometry (HPLCESI-MS). The results showed that the decomposition of L-aspartic acid considerably decreased in the presence of clay thus highlighting the protector role of clays and favors the stability of organic matter even under the possible high energy conditions of primitive environments. The principal product ofgamma radiolysis of L-aspartic acid was succinic acid produced by deamination reaction. On the other hand, when aspartic acid was irradiated in solid-state the main product was the L-aspartic acid dimer. Both radiolysis products are important for chemical evolution processes for L-aspartic acid in primitive environments.

Substitution of Aspartic Acid at Position 57 of the DQβ1 Affects Relapse of Autoimmune Pancreatitis

The existence of intracrystalline proteins and amino acids in calcium oxalate monohydrate was demonstrated by X-ray synchrotron diffraction studies. Their presence has implications for the destruction of calcium oxalate crystals formed in the urinary tract and the prevention of kidney stones. Although proteins are present in human kidney stones, their role in stone pathogenesis remains unknown. This investigation aimed to characterize the nature of the relationship between the organic and mineral phases in calcium oxalate monohydrate (COM) crystals grown in human urine and in aqueous solutions of proteins and amino acids to clarify the function of proteins in urolithiasis. COM crystals were grown in human urine and in aqueous solutions containing either human prothrombin (PT), Tamm-Horsfall glycoprotein (THG), aspartic acid (Asp), aspartic acid dimer (AspAsp), glutamic acid (Glu), glutamic acid dimer (GluGlu), or gamma-carboxyglutamic acid (Gla). Controls consisted of COM crystals precipitated from pure inorganic solutions or from human urine that had been ultrafiltered to remove macromolecules. Synchrotron X-ray diffraction with Rietveld whole-pattern peak fitting and profile analysis was used to determine nonuniform crystal strain and crystallite size in polycrystalline samples. Crystals precipitated from ultrafiltered urine had lower nonuniform strain than those grown in urine or in aqueous PT solution. Nonuniform strain was much lower in crystals grown in distilled water or in the presence of THG. For the amino acids, the highest nonuniform strain was exhibited by crystals grown in Gla solution, followed by Glu. Crystallite size was inversely related to nonuniform strain, with the effect being significantly less for amino acids than for macromolecules. Selected proteins and amino acids associated with COM crystals are intracrystalline. Although their incorporation into the mineral bulk would be expected to affect the rate of crystal growth, they also have the potential to influence the phagocytosis and intracellular destruction of any crystals nucleated and trapped within the renal collecting system. Crystals impregnated with protein would be more susceptible to digestion by cellular proteases, which would provide access to the crystal core, thereby facilitating further proteolytic degradation and mineral dissolution. We therefore propose that intracrystalline proteins may constitute a natural form of defense against renal stone formation.

The task of this work was to investigate the extraction capacity of various calixarenes for free and esterified amino acids from aqueous acid phases. Furthermore, this method was applied to aqueous extracts of Helleborus purpurascens. Generally, it is known that calixarenes can be used as extractants for ammonium compounds due to π-cation and lone pair cation interactions. As first, tert-Butyl-calix[6]arene and derivatives thereof were used. They had already proven their worth in previous investigations. In addition, tert-Butyl-hexahomooxa-calix[3]arene was used also, which can also enter into lone pair cation interactions. In addition to these well-known calixarenes, new calixarenes were produced and tested. Based on the tert-Butyl-hexahomooxa-calix[3]arene, a phosphor(III)bridged derivative was prepared, combining the three aromatic hydroxyl groups to a phosphite. As a seldom-described class of calixarenes, tert-Butyl-hexahomoaza-calix[3]arene derivatives were used. The nitrogen analogues of tert-Butyl-hexahomooxa-calix[3]arene could be produced as N-benzyl derivatives. The structure of the esterified carboxymethylated derivative of N,N',N″-Tribenzyl-tert-Butyl-hexahomoaza-calix[3]arene could be verified by X-ray structure analysis. It crystallized as a partial cone. The extraction capacity of the described calixarenes was investigated for amino acids from aqueous acidic solutions into an organic phase. For the testing were chosen asparagine, aspartic acid, tyrosine, tryptophane, phenylalanine and pipecolinic acid and their methyl esters. The amino acids and their methyl esters were dissolved in water at different pH values. The calixarenes were dissolved in dichloromethane (DCM) or chloroform. After this preparation, the aqueous acidic amino acid solutions were mixed with the solutions and shaken intensively. In addition, blank values were determined by extracting the aqueous stock solutions of the amino acids and their methyl esters with pure solvents. To determine the extraction rate, the phases were separated and each analysed using GC-FID, partially GC-MS(EI). The evaluation is performed in two ways. On the one hand the depletion in the aqueous phase and on the other hand the content in the organic phase was determined.

The importance of vacteria in the cycling of carbon in the Pamlico River estuary was studied by measuring the rates of uptake of organic compounds. Our methods allowed analysis with the Michaelis—Menten kinetics equations, and both the rates of uptake of dissolved free amino acids (DFAA) and glucose as well as the percentage of carbon subsequently respired as CO2 were determined. In addition, the concentrations of the amino acids in the water were determined using ion exchange chromatography. Other tests included measurements of primary productivity and of the effects of the other amino acids in the water upon the uptake of one amino acid. There was considerable variation in the heterotrophic activity over time and distance probably caused by patchiness in distribution of plankton and dissolved compounds in the water. Although there is some competition between amino acids being taken up, the effect upon kinetics measurements is probably negligible. Tests made every 3 hr showed a coefficient of variability (CV) of the measured maximum velocity of uptake (Vmax) of aspartic acid to be only 26%, and a similar CV was found for daily samples. In several instances the uptake of one amino acid was found to be competitively inhibited by the presence of another amino acid, but the concentrations necessary to inhibit were far above natural concentrations and such effects are probably unimportant in nature. Mutual inhibition was found between the similar amino acid pairs glutamic acid and aspartic acid, threonine and serine, glycine and alanine, and leucine and alanine. Highest Vmax values were found during the summer months and early fall and ranged from a high of 69.42 mg C/1°hr for alanine in August to less than 0.20 mg C/1°hr for most of the substrates tested in the colder months. The Vmax values for glucose uptake (0.06 to 9.64 mg C/1°hr) indicate that this estuarine system is one of the most microbially—active environments tested. The DFAA were presented in the water at concentrations of from 10 to 30 mg C/1; over half of this was ornithine, glycine, and serine. The DFAA were only about 0.2% of the total dissolved organic carbon in the water. Further, seasonal variations of DFAA concentrations, generally paralleling those of primary productivity, suggested that the amino acids originated from algal excretion and the decay of algal cells. The orders of abundance and concentrations of individual amino acids were similar to those reported for other bodies of water. When the natural concentration of a substrate is known the actual velocity of uptake (Vn) or flux for that substrate may be found. Flux rates were only 1%—10% of the Vmax values in the coldest months; the highest values were found in the warmest months. At each experimental concentration of amino acid, a certain amount was taken up, and a percentage of this amount was oxidized to caron dioxide. This percentage was constant for a particular amino acid in spite of varying experimental times, substrate concentrations, and temperatures. Leucine had the lowest percent respired (13%) while aspartic and glutanic acids had the highest (50%). Failure to correct uptake data for this respiratory loss introduces significant underestimation. The production of particulate material was calculated by correcting total uptake figures for each amino acid by its characteristic respiration percentage. Over 60% of the particulate production from amino acids was by uptake of alanine, leucine, valine, serine, glycine, aspartic acid, and glutamic acid. Such particulate production averaged 0.79 mg C/1.hr for the year and ranged from 0..6 to 2.37 mg C/1.hr; this is about 10% of the rate of production by algae during the summer months. This amount of particulate organic material is a significant contribution to this estuarine food chain.

Chiral compounds that crystallize as conglomerates from racemic mixtures are desirable from the perspective of chiral resolution since they are relatively easy to resolve using, for instance, preferential crystallization. Here, a screening methodology to find conglomerate cocrystals is demonstrated using racemic praziquantel (rac-PZQ). Cocrystallization of rac-PZQ was performed using liquid-assisted grinding (LAG) with 20 potential achiral coformers. LAG screening results show that N-phenyl urea (PU) and 1,3-dimethylthiourea (DMTU) are the only two achiral coformers that form new cocrystal compounds with rac-PZQ. Single-crystal structure determination of cocrystals grown from solution shows that the rac-PZQ:PU cocrystal is a racemic cocrystal with space group P21/c, while the rac-PZQ:DMTU cocrystal is a conglomerate cocrystal with space group P21. Under gentle conditions, rac-PZQ:DMTU forms a true conglomerate cocrystal from solution. However, when grown from solution under higher supersaturations, the obtained single crystals of rac-PZQ:DMTU appear to be disordered racemic twin cocrystals: these contain alternating domains of (almost) pure enantiomers of PZQ that are related by inversion symmetry. The disordered racemic twin cocrystals probably form due to refreshment of the enantiomer at the crystal surface being diffusion-limited from the bulk solution to the boundary layer. Discovery of a conglomerate cocrystal form of PZQ opens up exciting possibilities for the optical resolution of PZQ by preferential crystallization, allowing for easy preparation of pure enantiomer formulations of this important API.