Plant tissue preparation

Kertesz S reported that blenders usually only part the middle lamella and may not effectively homogenize or extract plant tissues. We have observed that acetone powders of plant tissues, prepared by standard procedures using the Waring Blendor, contain many multicellular bits of tissue. In acetone powders of peppermint we have even observed intact oil glands, still filled with essential oil. Robin Hill stated in a 1961 lecture that spinach leaves are a superior tissue for enzyme isolation because they have strong middle lamellae, which promote cell breakage as opposed to cell-cell separation. 

Plant tissue preparation procedures for calorimetric analyses are often very simple excise a section of plant tissue and place it in the calorimeter ampule. However, some standard precautions must be observed and some tissues require special handling or treatment to ensure meaningful measurements. 

The endo-action of the K. marxianus PG was demonstrated by a extremely rapid attack on plant tissue. This activity appears to be at least equivalent to that of several commercial preparations used for separating plant cells for protoplast preparation (RMC, unpublished data). Most of the endo-PGs produced by plant pathogens and saprophytes have so far been reported to possess macerating activity. PG secreted by K. marxianus CCT 3172 also had a strong activity in reducing the viscosity of cocoa pulp. Cocoa pulp generally contains 1 - 1.5% (w/w) of pectin consisting of 68% esterification and 11.6% methoxyl content [18]. 

Application of Preparative Layer Chromatography for the Separation of Secondary Metabolites from Plant Tissues... 

Classical approaches to plant DNA isolation aim to produce large quantities of highly purified DNA. However, smaller quantities of crudely extracted plant DNA are often acceptable for PCR analysis. Another efficient method for preparation of plant DNA for PCR is a single-step protocol that involves heating a small amount of plant tissue in a simple solution. Several factors influence nucleic acid release from tissue salt, EDTA, pH, incubation time and temperature. These factors must be optimized for different sample substrates. EDTA in the sample solution binds the Mg + cofactor required by the Taq polymerase in the PCR, so the EDTA concentration in the solution, or the Mg + concentration in the PCR, must be carefully optimized. 

Cellulose is a high molecular weight polymer of D-glucose with fi( 1 -4)-glycosidic bonds, found in plant fibres it is the major component of most plant tissues. Starch is another common polysaccharide, containing two polymers of glucose, amylose and amylopectin. It was used in some paint preparations and in the production of paper. Acid treatment of starch produces dextrins, which are used as adhesives and additives in water colour paintings. 

Fig. 1. The main choices in preparing plant tissues for microanalysis. 

This technique is simple in basic principle. Material is first rapidly frozen to the temperature of liquid nitrogen. It is then fractured, cryo-planed to produce a flat surface for analysis, and transferred to the cryo-stage of an SEM. It is analyzed while still frozen, and thus ion movement during tissue preparation should be minimal. A more detailed scheme of a typical procedure (45,46) is given in Subheading 3.4.2.1. This is undoubtedly the most popular microanalytical method with plant scientists at present, and as Table 1 shows it has been applied to a wide range of tissues and research topics (46-53). Recent developments include a... 

Lazof DB, Goldsmith JKG, Rufty TW, Suggs C, Linton RW. The preparation of cryosections from plant tissue an alternative method appropriate for secondary ion mass spectrometry studies of nutrient tracers and trace metals. JMicrosc 1994 176 99-109. 

In essence, an enzyme-catalyzed equivalent exists for almost every type of chemically catalyzed reaction, and thousands of these have been documented in comprehensive monographs and reviews (9-26). Many reactions have been observed in relatively specialized areas, particularly with groups of organic compounds such as the steroids, other terpenoids, antibiotics, aromatics, and alkaloids. Specific chemical reactions have been accomplished with intact and growing microbial cells, with plant and mammalian tissue preparations, and with... 

All of the principles and ideas covered in the previous section may be translated directly to the use of microorganisms as tools in the production of compounds of plant biosynthetic or biodegradative importance. Just as one finds microbial systems to be of value in preparing metabolites in mammalian systems, it may be possible to use microbial transformations to prepare derivatives of alkaloids that might be found rarely or only in very small quantities in plants. In this way, abundant prototype alkaloids may be used as microbial transformation substrates to provide a range of metabolites. As in the mammalian case, metabolism studies using plant tissues, tissue cultures, or cell-free extracts may be conducted in parallel with microbial metabolic systems. Metabolites common to both would be prepared in quantity by relatively simple fermentation scale-up methods. 

About 120 chemical constituents have been identified in chamomile as secondary metabolites, including 28 terpenoids, 36 flavonoids and 52 additional compounds [4]. A substantial part of drag effects are determined by the essential oil content. Oil is collected from flower heads, either by steam distillation or solvent extraction, for yields of 0.24-1.90% of fresh or dry plant tissue. Among the essential oil constituents the most active are /-/-a-bisabolol and chamazulene. /-/-a-bisabolol has demonstrated anti-inflammatory, antispasmodic, antimicrobial, antiulcer, sedative and CNS activity. Chamazulene is also anti-inflammatory. Topical applications of chamomile preparation have shown benefit in the treatment of eczema, dermatitis and ulceration [5]. 

Preparation of Other Antigenic Materials. Cotton plant tissues (stem, leaf, burr), cotton gin trash, baled cotton, clean cotton lint, both hand picked in the field and from plants grown in the greenhouse, cottonseed proteins, cottonseed hulls, house dust, and flax, soft hemp, sisal, and jute fibers, were extracted with deionized water. The purification process was, however, stopped to correspond to f-3 (see Figure 1). 

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