Iodine-amylopectin complex

Starch can form an intense, brilliant, dark blue-, or violet-colored complex with iodine. The straight chain component of starch, the amylose, gives a blue color while the branched component, the amylopectin, yields a purple color. In the presence of iodine, the amylose forms helixes inside of which the iodine molecules assemble as long polyiodide chains. The helix-forming branches of amylopectin are much shorter than those of amylose. Therefore, the polyiodide chains are also much shorter in the amylopectin-iodine complex than in the amylose-iodine complex. The result is a different color (purple). When starch is hydrolyzed and broken down to small carbohydrate units, the iodine will not give a dark blue (or purple) color. The iodine test is used in this experiment to indicate the completion of the hydrolysis. 

OD660 for Amylose/ Iodine complex OD530 for Amylopectin/ Iodine complex... 

Branching enzyme is assayed by measurement of the decrease in optical density of the amylopectin-iodine complex at 570 mu, after incubation at 30° and pH 7. Plant Q-enzymes have no effect on amylopectin and thus differ significantly from the animal branching-enzjrmes. For this reason, it was suggested " that the term Q-enzyme be confined to plant branching-enzymes. 

Determination of Kn 5 and Ki A plot of concen-tration of iodlne(I ) vs absorbance (640 nm) in the case of the amylose-iodine-complex/amylopectin-iodine-complex gives a typical rectangular hyperbola curve ( ), most often encountered in the field of enzyme kinetics. Similarly, the behaviour of many physical and biological systems can be described in... 

Amylose [9005-82-7] (C Hio05) (/br use in iodine complex formation). Amylopectin was removed from impure amylose by dispersing in aqueous 15% pyridine at 80-90 (concn 0.6-0.7%) and leaving the soln stand at 44-45° for 7 days. The ppte was re-dispersed and recrystd during 5 days. After a further dispersion in 15% pyridine, it was cooled to 45°, allowed to stand at this temperature for 12hours, then eooled... 

Tapioca and maize amylopectins have been sub-fractionated by fractional precipitation from aqueous solution with increasing amounts of methanol,64 71 and potato amylopectin by preferential precipitation on electrodialysis of the iodine complex.72 When these three amylopectins were subjected to chromatography, and eluted with a neutral buffer, all were found to consist of several sub-fractions.70... 

To determine the amylose content of starch, the iodine reaction has been most commonly used because amylose and amylopectin have different abilities to bind iodine. The methods such as blue value (absorbance at 680 nm for starch-iodine complex using amylose and amylopectin standards), and potentiometric and amperometric titration have been used for more than 50 years. These procedures are based on the capacity of amylose to form helical inclusion complexes with iodine, which display a blue color characterized by a maximum absorption wavelength (kmax) above 620 nm. During the titration of starch with iodine solution, the amount (mg) of iodine bound to 100 mg of starch is determined. The value is defined as iodine-binding capacity or iodine affinity (lA). The amylose content is based on the iodine affinity of starch vs. purified linear fraction from the standard 100 mg pure linear amylose fraction has an iodine affinity of 19.5-21.0mg depending on amylose source. Amylopectin binds 0-1.2mg iodine per 100mg (Banks and Greenwood, 1975). The amylose content determined by potentiometric titration is considered an absolute amylose content if the sample is defatted before analysis. 

Starch in the double mutant du wx is essentially 100% amylopectin thus, the wx mutant is epistatic to du. The absorbance maximum and extinction coefficient of the du wx branched polysaccharide-iodine complex are the same as for wx (Table... 

The mature kernel phenotype for the triple mutant ae du su is similar to that for su (Table 3.3). Sugar concentrations of mature272 and immature270 ae du su kernels are higher than those of either of the single mutants or the double mutants ae du or ae su, while starch concentration is similar to that in su and the two double mutants. The amylose percentage is close to that of normal when measured by either the blue value test (Table 3.6) or SEC (Table 3.7). However, in contrast to normal, a major proportion of the branched polysaccharide is smaller than typical amylopectin (as is that of ae su), and it elutes from an SEC column at an intermediate position (Table 3.7). The absorbance maximum and absorptivity (extinction coefficient) of the branched polysaccharide-iodine complexes are similar to those for ae and ae su, and are characteristic of loosely-branched polymers. The absorbance maximum of the amylose-iodine complex is similar to that of normal, du, or su, but the absorptivity is lower than for either (Table 3.7). No short chain amylose has been found in ae du su starch.90... 

Several assays (Fig. 1) are available to measure branching enzyme activity. The iodine assay is based on the decrease in absorbance of the glucan-iodine complex (Krisman, 1962) resulting from the branching of amylose or amylopectin by the enzyme, provided that a-amylases are absent from the enzyme preparation or their activity is greatly reduced by adjusting the assay conditions. 

The pathway for the formation of the amylose-iodine complex is well recognized, at least when aqueous solutions of KI and iodine are used. Relevant studies are lacking on the formation of the complexes between iodine vapor and both amylose and amylopectin. 

Mokhnach and Rusakova11 have shown that amylose-iodine and also starch-iodine complexes absorb at 226, 288-290, 344-360, and 585-620 nm. The first band appears only when iodine is added to the solution simultaneously with KI, and it is absent when only iodine is added. The longest-wavelength absorption may be indicative of the molecular size of the carbohydrate portion of the complex, as demonstrated106 in Table VIII. Spectra of iodine-amylopectin complexes were measured by Archibald et al.146 The UV absorption spectrum of the starch-iodine complex is shown... 

A method of determining airborne iodine has also been reported.241 Here, iodine is absorbed into 5% aqueous KI and spectrophotometrically determined at 590 nm in the form of its complex with starch. This method is selective with respect to bromine and chlorine, and the sensitivity of this method is 0.25 mg of I2 per m3 of air. The concentration of the, 31I isotope in water can be determined by a method involving isotope exchange in the starch-iodine complex.242 Flow-injection determination of ascorbic acid (0.1-40 mg/mL) has been proposed.243 Iodine is generated in the flow system as I3- ions, which are in turn exposed to starch to produce a steady signal at 350 and 580 nm. Ascorbic acid provides inversed maxima which are measured. This method is recommended for analysis of ascorbic acid in fruit juice, jam, and vitamin-C preparations. Use of the blue complex has also been reported for determination of sodium dichloro-isocyanurate in air.244 Obviously the blue reaction is applicable in the determination of amylose, amylopectin, and starch,245-252 as well as modified starches.245,253-255... 

Fig. 2.—Absorption Spectra of Polysaccharide-Iodine Complexes. (I) Mytiltis edulis glycogen, (II) rabbit liver glycogen, (III) waxy-maize starch (amylopectin). [Solutions contained 0.01% of polysaccharide and 0.02% of iodine in 0.2% of potassium iodide, and were read against an iodine—potassium iodide reference solution.]...

Methods of assay (m) = methylation (p) = periodate oxidation (e) = enzymic. B.V. = Blue Value (compare Refs. 3 and 4). Xmax = wavelength of maximum absorption of polysaccharide-iodine complex. Priming activity toward P-enzyme soluble starch = 1.0. Priming activity toward muscle phosphorylase glycogen = 100, corn amylopectin = 63. Molecular weight, 64,000 (compared with about 100,000 for the parent amylose). 

Only one case of Type IV disease, amylopectinosis, has been reported the storage polysaccharide had the properties of an amylopectin rather than a glycogen (for example, CL 21.2, X ax of the iodine complex 530 mju), and was only sparingly soluble in water, so that its presence in the liver stimulated the formation of fibrous tissue in that organ. A partial deficiency of branching enz3me was, therefore, indicated. 

Branching enzyme (BE) can be assayed in numerous ways. The iodine assay is based on the decrease in absorbance of the polysaccharide—iodine complex resulting from the branching of either amylose or amylopec-tin. Aliquots are taken at intervals during the incubation of the amylose or amylopectin with BE, and iodine reagent is added. The decrease of absorbance is measured at 660 nm for amylose and, for amylopectin, at 530nm. A unit of activity is defined as decrease in absorbance of 1.0 per min at 30 °C at the defined wavelength. 

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