Amylose separation

As mentioned previously, cellulosic phases as well as amylosic phases have also been used extensively for enantiomeric separations more recently (89,90). Most of the work ia this area has been with various derivatives of the native carbohydrate. The enantioresolving abiUties of the derivatized cellulosic and amylosic phases are reported to be very dependent on the types of substituents on the aromatic moieties that are appended onto the native carbohydrate (91). Table 3 fists some of the cellulosic and amylosic derivatives that have been used. These columns are available through Chiral Technologies, Inc. and J. T. Baker, Inc. 

An hplc assay was developed suitable for the analysis of enantiomers of ketoprofen (KT), a 2-arylpropionic acid nonsteroidal antiinflammatory dmg (NSAID), in plasma and urine (59). Following the addition of racemic fenprofen as internal standard (IS), plasma containing the KT enantiomers and IS was extracted by Hquid-Hquid extraction at an acidic pH. After evaporation of the organic layer, the dmg and IS were reconstituted in the mobile phase and injected onto the hplc column. The enantiomers were separated at ambient temperature on a commercially available 250 x 4.6 mm amylose carbamate-packed chiral column (chiral AD) with hexane—isopropyl alcohol—trifluoroacetic acid (80 19.9 0.1) as the mobile phase pumped at 1.0 mL/min. The enantiomers of KT were quantified by uv detection with the wavelength set at 254 nm. The assay allows direct quantitation of KT enantiomers in clinical studies in human plasma and urine after adrninistration of therapeutic doses. 

FIGURE 16.7 Native starch ( ) and fractions of native starch differing in their branching characteristics (nb/lcb amylose -type fraction scb amylopectin -t/pe fraction ) separated on semipreparative... 

FIGURE 16.12 Enzymatically synthesized amylose"-type nb/lcb glucans ( ) with a significant amount of the substrate glucose-1-PO4 separated on Sephacryl S-SOO/S-IOOO (60 + 9S x 1.6 cm) 3-ml fractions were collected for further analysis normalized (area = 1.0) eluogram profiles (ev) constructed from an off-line determined mass of carbohydrates for each of the pooled fractions flow rate 0.42 ml/ min V,xd = 126 ml, V , = 273 ml eluent O.OOS M NaOH. 

Examples with other Pirkle-type CSPs have also been described [139, 140]. In relation to polysaccharides coated onto silica gel, they have shown long-term stability in this operation mode [141, 142], and thus are also potentially good chiral selectors for preparative SFC [21]. In that context, the separation of racemic gliben-clamide analogues (7, Fig. 1-3) on cellulose- and amylose-derived CSPs was described [143]. 

Starches can be separated into two major components, amylose and amylopectin, which exist in different proportions in various plants. Amylose, which is a straight-chain compound and is abundant in potato starch, gives a blue colour with iodine and the chain assumes a spiral form. Amylopectin, which has a branched-chain structure, forms a red-purple product, probably by adsorption. 

Ferretti et al. (1988) used an amino column coupled to a derivatized amylose column (Chiralpak AS) operated in the reverse-phase mode to separate the enantiomers of the antifungal agent voriconazole from several chiral impurities and one achiral impurity. Three of the chiral impurities are the other enantiomer and corresponding diastereomers of voriconazole. More chiral impurities result from a chlorinated voriconazole. Additionally, this multidimensional method could baseline separate all but two of the chiral impurities into their respective enantiomers. These separations are shown in Figure 14.5. 

Aboul-Enein and Ali [78] compared the chiral resolution of miconazole and two other azole compounds by high performance liquid chromatography using normal-phase amylose chiral stationary phases. The resolution of the enantiomers of ( )-econazole, ( )-miconazole, and (i)-sulconazole was achieved on different normal-phase chiral amylose columns, Chiralpak AD, AS, and AR. The mobile phase used was hexane-isopropanol-diethylamine (400 99 1). The flow rates of the mobile phase used were 0.50 and 1 mL/min. The separation factor (a) values for the resolved enantiomers of econazole, miconazole, and sulconazole in the chiral phases were in the range 1.63-1.04 the resolution factors Rs values varied from 5.68 to 0.32. 

Perhaps the most important advance in the chemistry of starch was the realization of the apparent inhomogeneity of the material, followed by its fractionation into simpler components—first carried out quantitatively by Schoch1 in 1941. Starches can, in general, be separated into at least two chemically distinguishable entities amylose, a mixture of essentially unbranched chains, and amylopectin, a mixture of highly branched chains.2... 

The labile nature of the components necessitates that, for fundamental investigations, the starch should preferably be extracted from its botanical source, in the laboratory, under the mildest possible conditions.26 Industrial samples of unknown origin and treatment should not be used. The characterization of the starch would appear to entail (1) dissolution of the granule without degradation, (2) fractionation without degradation, (3) complete analysis of the finer details of structure of the separated components (including the possibilities of intermediate structures between the extremes of amylose and amylopectin), and (4) the estimation of the size, shape, and molecular-weight distribution of these fractions. 

Meyer31 has separated maize and potato amyloses into two fractions each, by (1) aqueous leaching, followed by (2) dissolution and precipitation of the remainder. His results of colorimetric, end-group determinations of size were ... 

Although it has been found that the separated amylose component can be readily orientated to yield fiber patterns, amylopectin usually gives poor or amorphous patterns. In the granule, however, amylopectin does exhibit crystallinity, since waxy maize starch gives a diffraction pattern and other waxy starches behave similarly.193 -195 (This suggests that the branch points in the amylopectin molecule may be in the amorphous part of the granule.)... 

Starch is the major energy store of plants chemically it is a polymer of glucose and occurs in two separate forms, amylose and amylopectin. The ratio of the two types depends on the plant that the starch has come from typically starch is 20 30% amylose and 70-80% amylopectin but there are amylomaizes with more than 50% amylose while waxy maize produces almost pure amylopectin with less than 3% amylose. 

The variation between the starch from different plants is considerable. The percentage of amylose varies from 27% in maize starch through 22% in potato starch to 17% in tapioca starch. The waxy maizes are unusual in that they are almost pure amylopectin. This is extremely convenient because it avoids the need to separate amylopectin from amylose chemically. 

The enantiomers of many compounds are eluted in reverse order on the 3,5-dimethylphenylcarbamates of cellulose (23x) and amylose (24n), suggesting that these two CSPs may be complementary in recognizing chirality. Many enantiomers unresolved on 23x can be resolved on 24n, and vice versa. Consequently, when two 23x and 24n columns were used for the resolution of 500 racemates, nearly 80% of the racemates were separated into enantiomers at least on either of 23x and 24n. 

Amylose inclusion compounds, 14 168 Amylosic phases, for chiral separations, 6 88-89... 

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