Use of glass fiber filters for the rapid preparation of in vivo absorption spectra of photosynthetic bacteria.

Usually, in vivo light absorption spectra of photosynthetic bacteria are obtained by measuring cell suspensions in glass or quartz cuvettes. In such arrangements, however, the scattering of light by the suspensions as well as sedimentation or even swarming behavior of motile cells (N. Pfennig, Arch. Mikrobiol. 42:90, 1962) may give rise to various errors. The error caused by light scattering in thin suspensions is partly eliminated by use of an opal glass behind reference and sample cuvettes (K. Shibata, A. A. Benson, and M. Calvin, Biochim. Biophys. Acta 15:461, 1954). C. S. Yentsch (Nature 179:1302, 1957) introduced a new technique for the estimation of chlorophyll concentrations in algal cultures. Algal cells are concentrated on a membrane filter, dried, and cleared with cedar oil. The cleared filter is then mounted in the spectrophotometer housing and absorbancy of the chlorophyll band at 670 m,u is measured. Air is the reference. Modification of this method with a blank filter as reference proved useful in measurements of visible light absorption by particulate matter in the oceans (C. S. Yentsch, Limnol. Oceanog. 7:207, 1962). Applying the same method for in vivo measurements of enrichment and pure cultures of photosynthetic bacteria, we found it useful for Chlorobacteriaceae, i.e., photosynthetic bacteria containing bacteriochlorophylls c or d (A. Jensen, 0. Aasmundrud, and K. E. Eimhjellen, Biochim. Biophys. Acta 88:466, 1964). At wavelengths above 750 mu, i.e., mainly in the in vivo absorption range of bacteriochlorophylls a and b, however, we were unable to use membrane-type filters because of their high apparent absorbancy in the near infrared region. This difficulty is now overcome by replacing the membrane with glass-fiber filters. By use of the method described below, the preparation of in vivo absorption spectra of photosynthetic bacteria representing all four bacteriochlorophyll types is possible in a time of 3 min per sample. The method proved to be useful