James H. C. Smith

Chlorophylls: Analysis in Plant Materials

Chlorophylls are the green pigments in plants some of which, and perhaps all, participate in photosynthesis. An understanding of their nature, genesis, transformation, and function in plants requires that they and their near chemical relatives be recognized qualitatively and oftentimes be determined quantitatively. In addition to the use of analytical procedures for the identification and determination of the chlorophylls as participants in plant physiological processes, these methods may be used in various other fields of science and technology some of which are genetics, plant nutrition, certain aspects of pure chemistry, crop production of land and sea, medicine, pharmaceuticals, food technology, and paleontology.

The action spectrum for the transformation of protochlorophyll to chlorophyll a in normal and albino corn seedlings.

Abstract 1. 1. A monochromator has been developed for use with a high intensity lamp by means of which leaves can be illuminated throughout the visible spectrum with light monochromatic within 5 mμ. The beam is of such intensity that the protochlorophyll-chlorophyll transformation produced by periods of illumination of from 20 sec. to 6 min. is sufficient for accurate measurement. 2. 2. Methods of analysis have been worked out whereby the percentage transformation of protochlorophyll can be determined with a precision of about 2%. 3. 3. An albino mutant of corn has become available which when grown in the dark contains approximately the normal amount of protochlorophyll, and upon illumination transforms it into chlorophyll. The action spectrum for the transformation has been measured in this albino strain and in the normal strain from which it segregates. 4. 4. The two corn strains and also bean leaves show a prominent peak in the action spectrum at 650 mμ. At this wavelength 20% of the protochlorophyll was transformed to chlorophyll in 18.4 sec. by an intensity of 79.8 ergs/cm. 2 /sec. 5. 5. Another prominent peak lies at 445 mμ. In normal corn and bean leaves, this peak is considerably lower than the peak at 650 mμ, the ratio being 0.66. In contrast, the albino corn uses blue light even more effectively than it does red, the ratio of the peaks at 445 and 650 mμ being 1.89. 6. 6. Since the normal corn leaves contain abundant carotenoids and the albino does not, the lower efficiency of normal etiolated leaves in the blue can be attributed to competitive absorption of light by carotenoid pigments. 7. 7. If some chlorophyll is formed by a pre-exposure to light and the leaves are then further illuminated at 680 mμ (light which is strongly absorbed by chlorophyll but only weakly by protochlorophyll) only a little additional chlorophyll is produced. The increase in chlorophyll can be attributed to the slight absorption by protochlorophyll. 8. 8. Because of the sensitivity and precision of the methods used, the action spectrum for the conversion of protochlorophyll in the leaf is known with an accuracy approaching that of the absorption spectrum of extracted protochlorophyll. The two spectra are similar and the positions of the spectra are in agreement if allowance is made for the shift due to the influence of solvent.

A Spectrophotometer accessory for measuring absorption spectra of light-scattering samples: Spectra of dark-grown albino leaves and of adsorbed chlorophylls

Abstract The opal-glass technique provides a means for obtaining serviceable absorption spectra of pigments in highly scattering samples. A convenient apparatus for using this technique in conjunction with a Beckman DK-2 spectrophotometer for routine spectrophotometric analysis of thin solid samples is described. Examples of absorption spectra obtained by use of this apparatus are given, namely, the changes of absorption that take place in dark-grown albino corn leaves when they are illuminated and the absorption spectra of pure chlorophylls a and b adsorbed on filter paper.

The Measurement of Artificial Radioactivity in Liquid Tracer Samples Using C11

Using C11 as tracer for the study of carbon dioxide absorption in plants, unexpected analytical difficulties were encountered. However, a satisfactory technique was developed for the rapid measurement of radioactivity in liquid samples. A degree of precision was obtained so that after the elapse of 6 half‐lives (2 hours) the sum of the activities taken from the reaction system was accurate to 1 or 2 percent. This required the determination of an accurate half‐life value for C11 (20.35 min.). The experimental procedures are described.