Pectic acid, determination

Another method is based on the same principle,112 in which the [14C]labelled methyl ester of D-galacturonan is prepared by esterification of pectic acid with [,4C]diazomethane. In the course of the enzymic de-esterification, aliquots are removed, and the unreacted substrate is precipitated with acidified ethanol or 1-propanol. After centrifugation, the labelled methanol in the supernatant liquor is determined in a liquid scintillation counter. An advantage of this method lies in the possibility of using, as substrates, short-chain oligo-D-galactosiduronates partially esterified with [14C]methanol. These substrates, beginning with the trisaccharide, are not soluble in 1 4 80% phenol-diethyl ether, which is used for the extraction of enzymically released, labelled methanol. 

Fig. 7. —Isolation of Endo-D-galacturonanase on a Column of Cross-linked Pectic Acid.210 [The Column (1.4 cm X 12 cm) was equilibrated with 0.1 M acetate buffer (pH 4.2). The first peak was eluted with the equilibrating buffer. Endo-D-galacturonanase was eluted with 0.1 M acetate buffer (pH 6.0). —, D-galacturonanase activity in 1 ml of eluate per min, determined by Somogyi reagent. The activity is expressed as change in absorbance at 530 nm, —, absorbance at 280 nm.]...

A series of O-acetyl derivatives of pectic acid has been prepared to study the intramolecular binding of Ca + ions to pectic acids. The electrostatic free enthalpy of dissociation of these polyacids and the rate of exchange of the counter-ion from Ca + to K+ were determined by potentiometric titration measurements. The binding of Ca + ions was characterized by the production of single ion affinity coefficients determined in solutions of the calcium salts of 0-acetyl pectic acids and by the circular dichroism measurements of calcium and potassium salts of the pectate derivatives. The intramolecular binding of Ca + ions to isolated pectate macromolecules is purely of electrostatic character in contrast to the intermolecular chelate binding of Ca + ions which takes place when the pectate macromolecules aggregate. 

The various groups of pectic substances are usually characterized by their solubility behavior. Polygalacturonides, like other polyuronides, can be quantitatively estimated by the amount of carbon dioxide formed upon heating with hot acid. Pectic substances of all sorts have been commonly determined by the formation and weighing of water-insoluble pectic acid or calcium pectate but unfortunately it is difficult to get these precipitates free from impurities (particularly other polysaccharides) present in the material analyzed. Today, the most common method of estimation is the spectrophotometric measurement of the colored complex formed upon the addition of alcoholic carbazole to the material to be analyzed after alkali treatment and heating with concentrated sulfuric acid. ... 

Rha, Ara and Gal are the neutral sugar components from all the fractions. Xyl is not present in Fla and is significantly present in the hemicellulose fractions, indicating that this monosaccharide is component of hemicellulosic polymers. Chemical composition of the water fractions were determined (Table V). High protein contents and the presence of O-acetyl-groups were observed in four aqueous fractions. Neutral sugar and uronic acid composition points to inclusion of these polymers in the class of pectic polysaccharides. 

Leitao, M. C. A. Alarcao-Silva, M. L. Januario, M. I. N. Azinheira, H. G. Galacturonic acid in pectic substances of sunflower head residues quantitative determination by HPLC. Carbohydr. Polym.,26(1995)165-169. 

Determination of pectic structures by specific degradation of pectins, via a carbodiimide 556 mediated Lossen rearrangement on alkaline hydrolysis of galacturonic acid residues 555, was reported (equation 248). 

Direct titrimetric method is a classic method for DE determination (National Research Council, 1972). In this method, pectin is first washed by acidic alcohol to convert the free carboxyl groups to the protonated form and is then washed with alcohol and dried. The washed sample is then dissolved in water and titrated with standard base and acid. The uronic content and DE are calculated from the neutralization and saponification equivalents of the pectic carboxyl groups. The existence of acetyl groups in some pectin samples leads to an overestimate of DE in the titration method. In this case, the copper binding method is an alternative approach (Keijbets and Pilnik, 1974). In the copper binding method, the amounts of copper ions that bind to the pectin molecules before and after saponification are measured by the complexometry, the ratio of which gives the DE. 

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