Extraction from plant tissue

This unit is composed of three separate procedures describing proanthocyanidin extraction from plant tissue (see Basic Protocol 1), purification (see Basic Protocol 2), and subsequent analysis by reversed-phase HPLC (see Basic Protocol 3). These protocols have been developed and used for analysis of grape skins, grape berries, and grape seeds. Without modification, wine, apples, and pears have also been analyzed using these procedures. 

To analyze proanthocyanidins, this unit relies on the susceptibility of the proanthocyani-din interflavonoid bond to acid catalysis. Interflavonoid bonds have different susceptibilities to acid catalysis (Hemingway and McGraw, 1983 Beart et al., 1985). In general, and for procyanidins and prodelphinidins, subunits that are bonded through the benzylic position (C4) are susceptible to acid catalysis. Most natural proanthocyanidins extracted from plant tissue have this type of interflavonoid bond and are therefore suitable candidates for this analytical technique. 

No complete picture of biological materials can be given without reference to their original condition in their natural environment. Brief consideration will, therefore, be given to the biosynthesis of the hemicelluloses and to their relationship, in the plant, to cellulose and lignin. Since much of our present knowledge of the chemistry of the hemicelluloses is based upon examination of materials isolated by alkaline extraction from plant tissues which, in many cases, have been previously treated to remove lig-... 

Hemicelluloses have been defined by Norman " as those cell wall polysaccharides which may be extracted from plant tissues by treatment... 

A direct injection method was proposed for phenolic acid extracts from plant tissue or soil, based on CZE at pH higher than the pK of the acids. Tetradecyltrimethylammonium bromide was added to reverse the electroosmotic flow. LOD was 1-7 p.M for eight phenolic acids at pH 7.20. ... 

Indole-3-acetic acid (Fig. 4.16) may be extracted from plant tissue after first homogenizing a 0.1-g sample and extracting the plant tissue with 75mAf potassium phosphate dibasic. After extraction, the pH is adjusted to 2.7 with 2.8 M phosphoric acid and the sample is dialyzed with 15 mL 0.1M potassium phosphate dibasic. The pH adjustment is necessary to protonate the acetic acid functionality for good retention on the C-18 sorbent. Use a trifunctional sorbent that is resistant to acid hydrolysis. 

CWs may be extracted from plant tissue and be subjected to adsorption experiments (Bush and McColl, 1987 Allan and Jarrell, 1989 Richter and Dainty, 1990 Grignon and Sentenac, 1991). On the basis of such experiments, Rufyikiri et al. (2003) found the following sequence of affinities of ions for CWs of banana root AP+ > H+ > Ca + > (Mg + or K+). Furthermore, CWs may be separated into fractions such as pectins, proteins, etc. prior to the experiments. For example. Franco et al. (2002) found the following sequence of affinities of ions for certain demethylated pectins Fe + > AF+ > Cu2+ Mn2+ > Zn2+ Ca +. 

Hormones are produced naturally by plants, while plant growth regulators are applied to plants by humans. Plant growth regulators may be synthetic compounds (e.g., Cycocel) that mimic naturally occurring plant hormones, or they may be natural hormones that are extracted from plant tissue (e.g., indole-3-acetic acid). 

Alkaloids can be extracted from plant tissues using dilute hydrochloric acid. Why does the use of acid enhance their water solubility ... 

S. BoreUa, G. Menofi md M. Leuenberger. Scunple homogeneity md cellulose extraction from plant tissue for stable isotope jmcdyses, in Handbook of stable isotope analytical techniques, P.A. Groot, (Ed.), Voliune 1, pp. 507-522, Elsevier (2004). 

There is some uncertainty as to which assay gives the most reliable results in combination with extracts from plant tissues rich in phenolic substances. The influence of such substances can never be predicted. It is therefore imperative to minimize interaction of these substances and protein in the course of sample preparation. For a more detailed discussion of this problem, see Guttenberger et al. (1994). 

The approach we have found to be most useful for the isolation and purification of Type 1 or 2 proanthocyanidin polymers is extraction from plant tissue with acetone-water mixtures (25, 37), separation of the monomers and lower oligomers from the resulting aqueous solution with ethyl acetate, adsorption on Sephadex LH-20 of the aqueous solution diluted with an equal volume of methanol and washing with the same solvent to eliminate impurities. The polymer is then displaced with acetone-water to yield freeze-dried analytically pure material (25). In the case of procyanidins, the ethyl acetate fraction contains monomers (catechin or epicatechin), dimers, trimers, and some tetramers, whereas the LH-20 fractions contain tetramers, on up to genuinely polymeric species. 

Several of the hydroxyproline-rich glycoproteins extracted from plant tissues have carbohydrate-binding activity these glycoproteins have the characteristics of lectins. These lectins or lectin-like glycoproteins have compositions which are similar to that of the cell wall hydroxyproline-rich glycoprotein. 

Various equations for the determination of total chlorophyll and individual amounts of chlorophylls a and b in extracts from plant tissues exist (see Holden 1976) and some of them (e.g. Amon 1949) have been widely used. Additional modifications to the equations have also been developed so as to permit an estimate of total carotenoids to be made from the spectrum of the same mixture in di-ethyl ether (Ziegler, Egle 1965 Gaudillere 1974). 

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