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An analytical methodology based on an O-[2-(methacryloyloxy)-ethylcarbamoyl]-10,11-dihydroquinidine (MQD)-silica hybrid monolithic column was developed for the enantioseparation of 9-fluorenylmethoxycarbonyl (FMOC) derivatized amino acids by nano-liquid chromatography. The mobile phase was optimized including the apparent pH, content of ACN, and concentration of the buffer to obtain a satisfactory enantioresolution performance. 27 FMOC derivatized amino acids including 19 protein and 8 non-protein amino acids were tested, and 19 out of them were enantiomerically discriminated obtaining baseline separation for 11 of them. Analytical characteristics of the method were evaluated for norvaline and tryptophan in terms of linearity, precision, accuracy, limits of detection (LOD) and quantitation (LOQ) showing good performance to be applied to the enantiomeric determination of these amino acids in dietary supplements. LOD and LOQ values were 9.3 and 31 μM for norvaline enantiomers and 7.5 and 25 μM for tryptophan enantiomers, respectively. The contents of d-norvaline and d-tryptophan were below their respective LODs in all the analyzed samples. Quantitation of l-tryptophan and l-norvaline showed good agreement with the labeled contents except for one sample which did not show presence of l-norvaline, contrary to the label indication., Graphical abstract Image 1, Highlights • A method was developed for the enantiomeric separation of amino acids by nano-LC. • A novel quinidine-silica hybrid monolith was employed as chiral column. • 19 protein and non-protein FMOC-amino acids were enantiomerically discriminated. • Analytical characteristics of the developed method were evaluated. • Norvaline and tryptophan were enantiomerically determined in dietary supplements.

An O-[2-(methacryloyloxy)-ethylcarbamoyl]-10,11-dihydroquinidine (MQD)-silica hybrid monolithic column was prepared by a facile "one-step" strategy within a 100 μm I.D. capillary. The influence of the methanol, ethylene glycol and water volume ratio, reaction temperature and time, cetyltrimethylammonium bromide and MQD monomers content and volume ratio of tetramethoxysilane and vinyltrimethoxysilane was investigated to obtain a satisfactory morphology of monolithic columns. The optimized MQD-silica hybrid monolithic column was evaluated in terms of permeability, stability, efficiency, reproducibility, and was characterized by scanning electron microscopy and nano-liquid chromatography. Among the 52 N-derivatized protein and non-protein amino acids, a total of 44 analytes could be baseline enantioseparated using the optimized conditions in either reversed phase mode (RPM) or polar organic phase mode (POM). The results showed that POM (ACN/MeOH/HAc/TEA (60/40/0.055/0.005, v/v/v/v)) offered better performance than RPM (10 mM ammonium acetate/ACN (30/70, v/v) (apparent pH=5.3)) in terms of enantioresolution and efficiency with shorter analysis times.

Chinoline alkaloids found in Cinchona bark still play an important role in medicine, for example as antimalarial and antiarrhythmic drugs. For the first time Supercritical Fluid Chromatography has been utilized for their separation. Six respective derivatives (dihydroquinidine, dihydroquinine, quinidine, quinine, cinchonine and cinchonidine) could be resolved in less than 7 min, and three of them quantified in crude plant extracts. The optimum stationary phase showed to be an Acquity UPC2 Torus DEA 1.7 μm column, the mobile phase comprised of CO2, acetonitrile, methanol and diethylamine. Method validation confirmed that the procedure is selective, accurate (recovery rates from 97.2% to 103.7%), precise (intra-day ≤2.2%, inter-day ≤3.0%) and linear (R2 ≥ 0.999); at 275 nm the observed detection limits were always below 2.5 μg/ml. In all of the samples analyzed cinchonine dominated (1.87%–2.30%), followed by quinine and cinchonidine. Their total content ranged from 4.75% to 5.20%. These values are in good agreement with published data, so that due to unmatched speed and environmental friendly character SFC is definitely an excellent alternative for the analysis of these important natural products.

Methods are presented to tritiate the plant natural product analogues dihydrocolforsin and dihydroquinidine.

A study of the osmium-catalyzed dihydroxylation of various olefins using air as the stoichiometric oxidant is described. Dihydroxylation takes place smoothly at an air pressure of 20 bar, at 50°C and pH 10.4. In the presence of dihydroquinine or dihydroquinidine derivatives (Sharpless ligands) asymmetric dihydroxylations occur with only slightly lower enantioselectivities compared to the classical K 3 [Fe(CN) 6 ] reoxidation system. In the case of stilbene the solvent system is crucial in determining the chemoselectivity of the reaction. The first example of a selective metal catalyzed oxidative cleavage of an olefin with air to give aldehydes is presented.

A non-aqueous capillary electrophoretic method developed for the enantioseparation of N-protected amino acids has been applied to the investigation of five new quinine and quinidine derivatives as chiral selectors: 1-adamantyl carbamoylated quinine, 3,4-dichlorophenyl carbamoylated quinidine, allyl carbamoylated dihydroquinine, allyl carbamoylated dihydroquinidine and 1-methyl quininium iodide. The composition of the background electrolyte was 12.5 mM ammonia, 100 mM octanoic acid in an ethanol-methanol (60:40 v/v) mixture containing a 10 mM concentration of the chiral selector. Under these conditions, the enantioseparation of a series of various benzoyl, 3,5-dinitrobenzoyl and 3,5-dinitrobenzyloxycarbonyl amino acid derivatives was studied with respect to selectand-selector relationship and enantioselectivity.

Dioxygen is able, under visible irradiation, to promote the high yielding and highly asymmetric dihydroxylation of α-methylstyrene in the presence of catalytic amounts of Os(VI), phthalazine dihydroquinidine chiral ligand [(DHQD) 2 PHAL].

Dihydroxylation of methyl ( E )-2,3-dideoxy-4,5:6,8-di- O -isopropylidene- l - xylo -oct-2-ene-4-ulofuranosonate ( 1 ) with osmium tetraoxide took place with high diastereoselectivity to give a 7:1 mixture of methyl 4,5:6,8-di- O -isopropylidene- α - l - glycero - d - galacto - ( 2 ) and - d - ido -oct-4-ulofuranosonate ( 3 ). When 1 was dihydroxylated in the presence of dihydroquinine and dihydroquinidine p -chlorobenzoate, an appreciable increase and decrease, respectively, in the 2 / 3 ratio was observed. Compound 2 was transformed into its 2,3-di- O -methyl derivative 4 which was deisopropylidenated to methyl 2,3-di- O -methyl- α - l - glycero - d - galacto -oct-4-ulopyranosonate ( 5 ) and subsequently degraded to dimethyl 2,3-di- O -methyl-(+)- l -tartrate ( 6 ). On the other hand, compounds 2 and 3 , separately, were isopropylidenated to the corresponding 2,3:4,5:6,8-tri- O -isopropylidene derivatives 7 and 8 , which were reduced with LiAlH 4 to the related 2,3:4,5:6,8-tri- O -isopropylidene- α - l - glycero - d - galacto - ( 9 ) and - d - ido -oct-4-ulofuranose ( 10 ). Finally, compounds 9 and 10 were deisopropylidenated to the corresponding l - glycero - d - galacto - ( 11 ) and l - glycero - d - ido -oct-4-ulose ( 12 ).

Stachysterone C was synthesized from 20-hydroxyecdysone (20-ECD). Sharpless asymmetric dihydroxylation of this rare ecdysteroid using osmium tetroxide and a chiral ligand afforded 24- epi -abutasterone, another rare ecdysteroid, and its C-24 epimer, abutasterone. High diastereomeric excess of the former ecdysteroid was obtained when the chiral ligands dihydroquinidine 1,4-phthalazinediyl diether and dihydroquinidine 2,5-diphenyl-4,6-pyrimidinediyl diether were employed. The two C-24 epimeric ecdysteroids exhibited similar moulting hormone activity in the Musca bioassay. However, they were significantly less active than 20-ECD.

Combination of variable temperature NMR and CD spectroscopy, and molecular mechanics computations shows that dihydroquinidine exists in two conformations , anti, open (80–90%) and syn, closed (10–20%) separated by a barrier of 8.3 kcal/mol in ether solutions .

The synthesis of a novel soluble polymer-bound ligand DHQD-PHAL-OPEG-OMe and its application in the catalytic asymmetric dihydroxylation (AD) reaction are described. 1,4-Dichlorophthalazine was used as the coupling reagent to connect dihydroquinidine and polyethylene glycol monomethyl ether (MW=5000, Fluka) while providing an aromatic group at the 9-O-position of dihydroquinidine. Enantiomeric excesses for trans-disubstituted olefins in the AD reaction, with K3Fe(CN)6 as secondary oxidant, are up to 98%.

The 22R,23R-homobrassinosteroid analogs are obtained in good yield from the corresponding precursor with a 22E-double bond by osmium-catalyzed asymmetric dihydroxylation using dihydroquinidine 9-O-(9′phenantryl) ether (DHQD PHN) as chiral agent.

Osmium catalyzed asymmetric dihydroxylation of alkenes has been reported using dihydroquinidine (DHQD) and dihydroquinine (DHQ) on a polystyrene support. Effect of other oxidants and additives has been examined.

Dihydroquinidine and dihydroquinine bisesters of malonic acid, adipic acid and terephthalic acids are used as chiral auxiliaries for osmium tetroxide catalysed asymmetric dihydroxylation of alkenes. Highly stereoselective osmylation of alkenes has been rationalized on the basis of π-π interaction of the alkenes with the ligand.

Catalytic asymmetric dihydroxylations of 1,3-enynes were studied using 9-O-(9′-phenanthryl) dihydroquinidine (PHN-DHQD) 1 and 1,4-bis-(9-O-dihydroquinidine) phthalazine (DHQD2-PHAL) 2. Terminal olefins showed moderate (38-79% ee) and trans-disubstituted olefins high enantioselectivities (73 – 97% ee).

Double asymmetric induction has been used to increase the diastereoselectivity of the dihydroxylation of the allylsilane +(11) using dihydroquinidine p-chlorobenzoate as catalyst.

A specific and sensitive assay for the separation and quantitation of quinidine, 3-hydroxyquinidine, quinidine-N-oxide, O-desmethylquinidine and dihydroquinidine is presented. The assay is shown to be sensitive to concentrations of 0.1 microgram/ml for all the above compounds when using a serum sample of 0.1 ml. The standard curve demonstrates linearity at concentrations from 0.1 to 5 micrograms/ml. The extraction procedure consists of adjusting the serum to an alkaline pH and extracting once with a mixture of methanol-dichloromethane (15:85). The organic extract is dried and the residue is solubilized in mobile phase. The chromatographic conditions are an isocratic delivery of the mobile phase 0.01 M K2HPO4-acetonitrile (96:4) through a C18 column at ambient temperature. Detection of the compounds of interest is by ultraviolet absorption at a wavelength of 210 nm. For each compound the inter-assay variation is less than 10% and the intra-assay variation is less than 15%. No interfering compounds were detected when a commercially prepared serum spiked with 28 commonly used therapeutic compounds was assayed by this method. The analytical method presented here for the isolation and quantitation of quinidine, several active metabolites, and dihydroquinidine has adequate sensitivity and specificity for monitoring the concentration of quinidine and quinidine metabolites in patient samples.

The highest enantioselectivities to date for various olefins are obtained in the osmium-catalyzed asymmetric dihydroxylation using new 9-O-aromatic dihydroquinidine and dihydroquinine ligands. A ligand structure-enantioselectivity relationship (LSER) is developed, and representative results for quinine-based ligands are reported for the first time.

The effects of dihydroquinidine on the metabolism of quinidine by 10,000 g supernatant fractions of rabbit and rat liver homogenates and on the disposition of quinidine in the rabbit were investigated. From Lineweaver-Burk plots, the following mean +/- S.E.M. values for V(max) and K(m) for quinidine and K(i) for dihydroquinidine for the rabbit liver preparations were obtained: 240 +/- 50 nmoles/min/g liver,0.55 +/- 0.14 mM, and 0.56 +/- 0.09 mM respectively. The corresponding values for the rat liver homogenates were 74 +/- 6 nmoles/min/g liver, 0.12 +/- 0.02 mM and 0.14 +/- 0.06 mM. From Dixon plots, the following values for K(i) for dihydroquinidine were obtained:0.54 +/- 0.09 mM for the rabbit liver preparations and 0.14 +/- 0.04 mM for the rat liver homogenates. In rabbits pretreated with dihydroquinidine for an average of 26 days, no changes in the distribution and elimination half-life values for quinidine, total body clearance, or apparent volume of drug distribution were observed when compared to the control quinidine disposition constants obtained in the same animals. As their structures suggested, the data showed that the interactions of dihydroquinidine and quinidine during drug metabolism by rabbit and rat liver 10,000 g supernatant fractions were competitive. Additionally, the affinities of dihydroquinidine and quinidine for the drug-metabolizing enzymes in these preparations were the same. The data also suggested that the small amounts of dihydroquinidine normally found in quinidine preparations probably have no significant effect on the disposition of quinidine in the body when therapeutic doses of the drug are used.

A rapid and specific high-performance liquid chromatographic (HPLC) assay was developed for the determination of the antiarrhythmic quinidine analog, 7'-trifluoromethyldihydrocinchonidine.2HCl ([I].2HCl) in plasma and urine. The overall recovery of [I] from plasma was 86 +/- 9% with a sensitivity limit of detection of 0.2 microgram/ml. The assay involves extraction of [I] into benzene--methylene chloride (9:1) from plasma or urine made alkaline with 0.1 N sodium hydroxide (pH 13) and saturated sodium chloride, the residue of which is dissolved in methylene chloride, an aliquot of which is analyzed by HPLC using adsorption chromatography on silica gel with UV detection at 254 nm. The mobile phase composed of methylene chloride--methanol--conc. ammonium hydroxide (95.5:4:0.5) yields baseline resolution of quinidine used as the internal (reference) standard, compound [I] and dihydroquinidine, a common contaminant in quinidine. The assay was applied to the analysis of plasma and urine samples taken from a dog administered a single 20 mg/kg dose via intravenous and oral routes. The stability of [I] in human plasma for up to 37 days of storage at -17 degrees C was also demonstrated.

The application of high-performance liquid chromatography to resolve the individual alkaloids present in marketed Cinchona alkaloids was investigated. Normal-phase and several reversed-phase systems were evaluated. The proposed procedure uses an alkylphenyl column; it adequately resolves quinidine, quinine, dihydroquinidine, dihydroquinine, cinchonine, cinchonidine, dihydrocinchonine and dihydrocinchonidine. Epiquinidine, epiquinine, quininone and quinitoxine are also resolved from quinidine and dihydroquinidine. This high degree of resolution enables the analysis of quinidine and its salts for their usual composition and establishes the absence of any cross-contamination or decomposition. The proposed procedure was applied to currently marketed samples of quinidine salts and their dosage forms. It was also applied to samples that were cross-contaminated or which contained decomposition products.

An high-performance liquid chromatographic assay for quinidine and dihydroquinidine sulphates in pharmaceutical dosage forms has been developed. The method makes use of a reversed-phase system with a C18 bonded column and theophylline as an internal standard. Recovery of both compounds is quantitative and the method compares favourably with that of the United States Pharmacopoeia with regard to speed and precision. The method should also be suitable for the control of quinine salts in pharmaceuticals.

Dihydroquinidine acetate promoted osmylation of acetals of E-α,β- unsaturated aldehydes occurs with up to 40:1 stereoselection to afford the corresponding protected syn -α,β -dihydroxy aldehydes.

A rapid, sensitive, accurate method for determination of quinidine in plasma has been developed using ion-pair extraction and high-performance liquid chromatography. The method, which is capable of distinguishing between quinidine and dihydroquinidine, involves acidification of plasma with perchloric acid, extraction with methyl isobutyl ketone and chromatography of the carbonate-washed extract on a silica gel column with a mobile phase of methylene chloride—hexane—methanol—perchloric acid (60:35:5.5:0.1) followed by fluorometric detection. The procedure is sensitive to below 50 ng/ml (coefficient of variation 6.6%) and compares favourably with a standard spectrofluorometric method when tested with plasma from volunteer subjects.

Liquid-liquid chromatography based on the ion-pair partition technique gives separation systems of high efficiency when silica micro-particles are used as the support for the stationary phase. With 10-μm particles, plate heights of the order of 40–70 μm have been achieved with a linear velocity of 0.25 cm/sec. The retention in ion-pair partition systems is determined by the nature and concentration of the counter ion, and the properties of the mobile phase also have a major influence. It is often possible to predict the selectivity, and this can be controlled by varying the composition of the mobile phase. This paper describes the application of ion-pair partition chromatography to the bioanalysis of drugs, drug metabolites and biogenic substances. Typical counter ions in the stationary phase were methanesulphonate and perchlorate for ammonium compounds and tetrabutylammonium for the separation of organic anions. Determinations by liquid chromatography were demonstrated for quinidine and dihydroquinidine, metanephrine and normetanephrine and for imipramine and its demethyl metabolite in plasma. A quaternary ammonium compound, QX-572, was determined in urine and chromatograms are shown for the isolation of indoleacetic and hydroxy-indoleacetic acid in urine. The methods have been used in routine analysis. Ultraviolet detection has permitted the determination of highly absorbing compounds down to the 10-ng level in plasma and urine.

A specific and precise assay, capable of quantitating in human plasma simultaneously but separately quinidine, dihydroquinidine and the quinidine metabolites 2'-quinidinone, 3-OH-quinidine and a third metabolite found--tentatively identified as the product formed by rearrangement of quinidine-N-oxide-is reported. The assay uses a normal phase high-performance liquid chromatographic (HPLC) system with a variable-wavelength UV detector at 235 nm and has a limit of sensitivity at approximately 20 ng/ml. The mobile phase consists of hexanes-ethanol-ethanolamine (91.5:8.47:0.03). A 2-ml plasma sample is worked up by adding primaquine base as an internal standard and extracting with ether-dichlormethane-isopropanol (6:4:1). The organic extract is evaporated and the residue reconstituted in 100-600 micron1 of mobile phase and an aliquot injected onto the column. Comparison of this procedure with the Edgar and Sokolow (dichloroethane) extraction--fluorescence procedure and with the Cramer and Isaksson (benzene) double extraction--fluorescence assay indicates that both fluorescence procedures give quinidine concentrations up to 2.3 times those determined by HPLC. These discrepancies were shown to be due to carry-over of metabolites and some extraneous background fluorescence.

Quinidine therapy is one of the most common causes of the acquired long QT syndrome and the morphologically distinctive tachyarrhythmia torsade de pointes. Clinical data from our institution and others have revealed a number of characteristic features: quinidine plasma concentrations are generally low, marked QRS prolongation is absent, hypokalemia is frequent and abrupt heart rate slowing just before the initiation of a paroxysm is almost invariable. The lack of correlation between plasma quinidine concentrations and this adverse drug effect raises the possibility either that external factors (for example, hypokalemia) modulate the response to quinidine in vivo or that one or more unmeasured active metabolites play a role. Therefore, the effects of alterations in extracellular potassium and stimulation rate on the electrophysiologic effects of quinidine were examined in canine Purkinje fibers. It was found that a form of triggered automaticity, early afterdepolarizations, is reliably produced in the presence of quinidine when extracellular potassium is lowered and the stimulation rate is slowed. More recently, the effects of a number of quinidine metabolites, as well as the commonly found impurity dihydroquinidine, were characterized in canine Purkinje fibers in a similar fashion. Although quinidine was the most potent of the substances tested, both dihydroquinidine and 3-hydroxyquinidine prolonged action potential and produced early afterdepolarizations as did quinidine at long cycle lengths. Quinidine-induced torsade de pointes is a potentially lethal adverse drug effect, occurring in 1 to 3% of patients. Hypokalemia and slow heart rates are commonly observed in a clinical setting and, in the tissue bath, quinidine and several of its metabolites induce abnormal automatic behavior when extracellular potassium is lowered and stimulation rate is slowed. It is concluded that early afterdepolarizations, a form of triggered automaticity, may be responsible for induction of quinidineinduced arrhythmias in humans.

Dihydroquinidine is a commonly encountered contaminant in quinidine raw materials. The USP allows 0-20% dihydroquinidine in quinidine products, but the assays used to quantitate dihydroquinidine have been lengthy or have required sophisticated equipment. The present method separates dihydroquinidine from quinidine and provides rapid, precise quantitation of both dihydroquinidine and quinidine. The clinical importance of dihydroquinidine contamination of quinidine dosage forms remains unanswered.

A high-pressure liquid chromatographic assay for quinidine and dihydroquinidine was developed. A cation-exchange column was utilized with an eluting solvent pH of about 9. The internal standard was cinchonine. The reproducibility and precision of this method were evaluated by analyzing replicate samples and by comparing results with those obtained from a TLC-fluorometric procedure. In addition, several drugs were evaluated to ascertain whether they interfered with the analysis of quinidine and dihydroquinidine.