**10. Chromatographic techniques as an aid in the study of the fine structure of pectic polysaccharides**

Chemical and enzymatic hydrolysis has been used to produce fragments of pectic polysaccharides in order to study the fine structure of pectins. The differences in the lability of glycosidic linkages to acid hydrolysis (see section 3.2) allowed the homogalacturonan region to

be isolated and its minimal length estimated. Under controlled conditions, acid hydrolysis of RG-I gives a fraction of intermediate molar mass that is rich in GalA and Rha. The neutral side chains are quickly hydrolysed to form low molar mass oligosaccharides [49; 42].

Characterization of Apple Pectin – A Chromatographic Approach 339

RG-II Rhamnogalacturonan II DHA Docosahexaenoic acid

DE Degree of methyl-esterification

XGA Xilogalacturonana ARA-I Arabinogalactan I ARA-II Arabinogalactan II API Apiogalacturonan

DA Degree of acetylation RI Differential refractometer LALLS Low angle laser light scattering MALLS Multi-angle laser light scattering

PC Paper chromatography GC Gas-liquid chromatography TLC Thin layer chromatography

AUA Uronic acids

Maria Helene Giovanetti Canteri

Alessandro Nogueira and Gilvan Wosiacki\*

*Federal University of Paraná, Curitiba, Paraná, Brazil* 

Carmen Lúcia de Oliveira Petkowicz

eucarpia2011.woiak.sggw.pl.

**Author details** 

**12. References** 

25-30.

Corresponding Author

 \*

KDO 3-Deoxy-D-manno-oct-2-ulosonic acid

HPSEC High Pressure Size Exclusion Chromatography

HPLC High performance liquid chromatography

FT-IR Fourier transform infrared spectroscopy

*Federal Technological University of Paraná, Ponta Grossa, Paraná, Brazil* 

GC-MS Gas chromatography associated to mass spectroscopy

pulsed amperometric detection

HPAEC-PAD High performance anion exchange chromatography coupled with

*State University of Ponta Grossa, Av. General Carlos Cavalcanti, Ponta Grossa, Paraná, Brazil* 

[1] Denardi F, Camilo AP. Novas cultivares de macieira: proposta de nova composição de pomares com polinizadoras/produtoras. Revista Agropecuária Catarinense, 1997; 10(2):

[2] Juniper BE, Watkins R, Harris SA. The origin of the apple. In: Eucarpia Symposium on Fruit Breeding and Genetics, XIII, Poland, Warsaw, 11-15 Sept. 2011. Available in

RG-II was first isolated from cell walls using enzymatic treatment. The structure of RG-II was investigated using a combination of enzymatic and controlled acid hydrolysis. Using chemical fragmentation, four oligosaccharides from side chains were obtained [42].

The products from chemical and enzymatic hydrolysis are further fractionated by chromatographic techniques and structurally characterized. Usually, ion-exchange and size exclusion chromatography are used to fractionate the segments of pectic chains produced in the partial hydrolysis. The recovery of oligosaccharides with low degree of polymerization greatly facilitates the structural studies. Elucidation of the sequence of glycosyl units with the methods current available is only possible for oligosaccharides. This approach allowed the identification of xylogalacturonan in the modified hairy region of apple pectin [50].

One of the steps in the structural analysis of pectins is the determination of the glycoside linkages. Methylation analysis is one of the main tools to determine glycosydic linkages of polysaccharides and their oligomers. In this procedure, the free hydroxyl groups of pectins or their oligosaccharides are methylated. Then, the partially methylated material is hydrolysed and submitted to reactions to afford volatile derivatives. The products from this procedure are separated and analyzed using the chromatographic method of GC-MS [51].
