Modern Methods of Plant Analysis When the handbook Modern Methods of Plant Analysis was first introduced in 1954 the considerations were: 1. the dependence of scientific progress in biology on the improvement of existing and the introduction of new methods; 2. the difficulty in finding many new analytical methods in specialized journals which are normally not accessible to experimental plant biologists; 3. the fact that in the methods sections of papers the description of methods is frequently so compact, or even sometimes so incomplete that it is difficult to reproduce experiments. These considerations still stand today. The series was highly successful, seven volumes appearing between 1956 and 1964. Since there is still today a demand for the old series, the publisher has decided to resume publication of Modern Methods of Plant Analysis. It is hoped that the New Series will be just as acceptable to those working in plant sciences and related fields as the early volumes undoubtedly were. It is difficult to single out the major reasons for success of any publication, but we believe that the methods published in the first series were up-to-date at the time and presented in a way that made description, as applied to plant material, complete in itself with little need to consult other publications. Contributing authors have attempted to follow these guidelines in this New Series of volumes.

Inhalt

The Use of Combined Gas Chromatography-Mass Spectrometry in the Analysis of Plant Growth Substances.- 1 Introduction.- 2 Identification by GC-MS.- 2.1 Derivatisation.- 2.1.1 Methylation.- 2.1.2 Trimethylsilylation.- 2.1.3 Permethylation.- 2.2 Gas Chromatography.- 2.3 Qualitative Mass Spectrometry.- 2.4 Quantitative Mass Spectrometry.- 3 Gibberellins.- 3.1 Extraction and Purification.- 3.2 GC-MS.- 4 Abscisic Acid and Related Compounds.- 4.1 Qualitative GC-MS.- 4.2 Quantitative GC-MS.- 5 Brassinosteroids.- References.- Applications of Mass Spectrometry for the Examination of Pectic Polysaccharides.- 1 Introduction.- 2 Structural Analysis of Pectic Polysaccharides.- 3 Determination of the Nature of the Glycosidic Linkages.- 4 Separation and Identification of Partially Methylated Alditol Acetates.- 5 Extensions of Methylation Analysis.- 5.1 Controlled Partial Acid Hydrolysis Studies.- 5.2 ?-Eliminative Degradation Studies.- 5.2.1 Neutral Glycosyl Residues Linked to Galacturonosyl Residues.- 5.2.2 Glycosyl Residues Linked to 0-4 of 2,4-Linked Rhamnopyranosyl Residues.- 6 Sequencing of Sugar Residues in Pectins.- 6.1 Partial Acid Hydrolysis.- 6.1.1 Characterisation of Oligosaccharides as Permethylated Derivatives.- 6.2 Partial Acetolysis.- 6.3 Enzymatic Hydrolysis.- 7 Sequencing of Pectic Polysaccharides by Partial Depolymerisation of Permethylated Derivatives.- 8 Experimental.- References.- GC-MS Methods for Cyclic Nucleotides in Higher Plants and for Free High Unsaturated Fatty Acids in Oils.- 1 Introduction.- 2 Methods of GC-MS.- 3 Adenosine-3?:5?-Monophosphate (cAMP) in Maize Seedlings (Zea mays).- 4 An Isotope Dilution GC-MS Spectrometric Assay for cAMP in Cultured Tobacco Tissue.- 5 Stability of Cyclic Purine Nucleotides in the Presence of Hydrochloric Acid During Extraction.- 6 Guanosine-3?:5?-Monophosphate (cGMP) in Maize Seedlings (Zea mays).- 7 GC-Separation of Synthetic cAMP and cGMP in a Mixture.- 8 Cyclic Pyrimidine Nucleotides in Plants?.- 9 Free High Unsaturated Fatty Acids in Oils.- 10 Conclusions.- References.- GC-MS Methods for Lower Plant Glycolipid Fatty Acids.- 1 Introduction.- 2 Extraction of the Plant Material.- 2.1 Handling and Storage.- 2.2 Extraction with Organic Solvents.- 2.3 Purification of the Extract.- 3 Separation of Glycolipids from the Total Lipid Extract.- 3.1 Column Chromatography.- 3.2 Thin-Layer Chromatography (TLC).- 3.3 Other Applications.- 4 Isolation of the Glycolipids.- 4.1 Thin-Layer Chromatography.- 4.2 Localization of the Glycolipids on TLC.- 4.3 Removing the Spots from the TLC Plates.- 5 Derivatization of the Glycolipid Fatty Acids for GC-MS.- 5.1 General Features.- 5.2 Formation of Methyl Esters.- 5.3 Silylation and Other Methods.- 6 GLC Instrumentation for Fatty Acid Analysis.- 6.1 General Features.- 6.2 Carrier Gas System.- 6.3 Injection and Injectors.- 6.4 The Detector.- 7 Column Selection for Fatty Acid GLC Analysis.- 7.1 Column Types.- 7.2 Supports, Liquid Phases, and Their Characteristics.- 8 Interpretation of GC Data and Calculation of Results.- 8.1 Identification of Peaks Using Standard Compounds.- 8.2 Quantitation of Results.- 8.3 External and Internal Standardization.- 9 Fatty Acid Ester Structure Determination by GC-MS.- 9.1 Equivalent Chain Lengths (ECL).- 9.2 Semilogarithmic Correlations.- 9.3 Mass Spectrometers and Their Function Principles.- 9.4 Interpretation of Mass Spectra of Fatty Acid Esters.- 10 Abbreviations.- References.- Analysis of Phospholipid Molecular Species by Gas Chromatography and Coupled Gas Chromatography-Mass Spectrometry.- 1 Introduction.- 2 Lipid Preparation.- 2.1 Lipid Extraction.- 2.2 Purification of Phospholipids.- 2.2.1 Column Chromatography.- 2.2.2 Thin Layer Chromatography.- 3 Formation of Derivatives for GC or GC-MS.- 3.1 Phospholipase C Treatment.- 3.2 Conversion of Diacylglycerols to Silyl Derivatives.- 3.2.1 Formation of Trimethylsilyl Derivatives.- 3.2.2 Formation of tert-Butyldimethylsilyl Derivatives.- 4 Gas Chromatography.- 5 Mass Spectrometry.- 5.1 Instrumentation.- 5.2 Operating Conditions.- 5.3 Identification of Molecular Species.- 5.4 Quantitation of Molecular Species by GC-MS.- 5.5 Quantitation of Molecular Species by GC-MS Following Reduction of Double Bonds Using Deuterium.- 5.6 Direct MS Analysis of Underivatized Phospholipids.- 6 Determination of Positional Distribution of Acyl Chains Using Phospholipase A2.- 7 Conclusion.- References.- GC-MS of Plant Sterol Analysis.- 1 Introduction.- 2 Development of GC-MS Plant Sterol Analysis.- 3 Operations Before GC-MS Sterol Analysis.- 3.1 Extraction and Isolation of Plant Sterols.- 3.2 Free Sterols and (or) Sterols from Steryl-Esters.- 3.3 Purification of Sterolic Fractions.- 3.4 Derivatization.- 4 Characterization of Sterols.- 4.1 Characterization of Sterols by GC Data.- 4.2 Characterization of Sterols by MS Data.- 4.3 Characterization of Sterols by GC and MS Data.- 4.3.1 A Typical Analysis of 4-Demethyl and 4,4-Dimethyl Sterols from Zea mays.- 4.3.2 Co-Occurrence of ?5- and ?7-Sterols in Tracheophytes.- 4.3.3 Side Chain-Hydroxylated Sterols from Red Algae.- 4.3.4 4-Methyl Sterols of Dinoflagellates.- 5 Conclusion.- References.- GC-MS Methods for Terpenoids.- 1 Introduction.- 2 Isolation Methods.- 3 Prefractionation and Ancillary Reactions.- 4 Gas Chromatography.- 5 Retention Data.- 6 Mass Spectrometry.- References.- GC-MS of Auxins.- 1 Introduction.- 2 The Compounds Involved.- 3 Reference Compounds.- 4 Extraction.- 5 Purification.- 6 Columns for GC.- 7 Injection Techniques.- 8 Derivatisation.- 9 Interface Between GC and MS.- 10 Mass Spectrometer.- 11 Data Systems.- 12 Ionization.- 13 GC-MS Strategy for Auxin Analysis.- 14 Quantification.- 15 The Internal Standard.- 16 Experimental Procedure.- 17 Conclusions.- References.- GC-MS Methods for the Quantitative Determination and Structural Characterization of Esters of Indole-3-Acetic Acid and myo-Inositol.- 1 Introduction.- 1.1 Discovery of IAA-Inositols.- 1.2 Occurrence of IAA Conjugates.- 1.3 Importance of Measuring and Identifying Hormone Conjugates.- 2 Quantitative Analysis and Identification of the IAA-Inositols.- 2.1 Analysis After Hydrolysis.- 2.1.1 Methods for Hydrolysis of IAA Conjugates.- 2.1.2 Use of Internal Standards.- 2.2 Analysis Before Hydrolysis.- 2.2.1 A Quantitative Estimation of IAA-Inositol Using [3H]-IAA-myo-Inositol as an Internal Standard.- 3 Qualitative Analysis of IAA-Inositols.- 3.1 The Inositol Moiety.- 3.2 Derivitization of IAA-Inositols for GC-MS.- 3.3 Mass Spectral Fragmentation Pattern.- 3.3.1 1-dl-1-O-(Indole-3-Acetyl)-my…