Hidrológiai Közlöny 1967 (47. évfolyam)
7. szám - A „Szervesanyag meghatározási problémák édesvizekben” című 1966. szeptember 25–28. között Tihanyben rendezett Szimpózium előadásai - Shapiro, Joseph: Különböző tavakból száramzó szervesnyagok összehasonlítása
290 Hidrológiai Közlöny 1967. 7. sz. Shapiro, J.: Különböző tavakból származó szervesanyagok The gel, which was swollen in 0.5 M aqueous ammónia before use, was supported in a "Sephadex" column 1.5 cm in diameter and 30 cm high. Portions of extracts weighing approximately 1 mg were dissolved in about 100 ?. of 0.5 M ammónia before being applied to the top of the column, and elution was performed with 0.5 M ammónia under slight head pressure. In order to achieve maximum sensitivity and aeeuracy the effluent was monitored at 350 m/t with a double beam spectrophotometer (Bausch & Lomb 'Spectronic 600') using a continuous flow cell with a capacity of 0.1 ml and a path length of 1 cm (Beckman). The absorbance was reeorded in linear fashion on a strip chart recorder (Sargent, model SRL). In this fashion quantitative results were obtained and many samples could be run with little effort. The results shown in the figures are direct tracings of the recorder charts. Choice oí Sephadex Povoledo and Gerletti [6] recommended Sephadex G-25 in their work on lacustrine materials, but Gjessing [7] had no success with either this grade or with G-50. Instead, Gjessing used Sephadex G-75, G-100, and G-200. The results of testing various grades of Sephadex under my conditions are shown in Figure 2. Clearly the best separations are obtained with G-50. G-10 excludes all of the color while G-75 retards all of the color. G-25 is somewhat intermediate, giving the same peaks as G-50 but less well separated. Numerous experiments confirm these results and show too that no further resolution is obtained with longer columns. Therefore all experiments reeorded were done with Sephadex G-50 (fine) using the column described above. As the rate of elution could not be exactly dupücated, the results are plotted, on the hasis of time, as "high" or "low" molecular weights, which they probably are, rather than as a function of elution volume. The particular laké extract used in the experiment shown in Figure 2 seems to have as many as 6 different components, although, as the elution volume of the first peak corresponds to Vo, or the void volume, it is possible that there are even more. In order to show that the separations are valid i.e. that the components are not the result of sample preparation or operational procedure, a series of experiments was done as follows: Eífect oí Drying and Heating Figure 3 shows elution patterns of samples wlűch were either not heated and dried (i. e. extracted into butanol from the laké water concentrate and back extracted into ammónia solution), or dried at 40°C or 80°C. In this particular case drying, even at 80°C, seems to have had no effect. Thus the fractions are not generated by drying. However in Figure 4 which shows the results of a similar experiment on an extract from a different laké, drying at 80°C seems to have significantly reduced the proportion of high molecular weight components. The reason that the 40°C fraction has a larger low molecular weight peak Molecular Weight Fig. 2. Elution patterns of the yellow acids from Oreen Laké, Minnesota, using the grades of Sephadex indieated Molecular Weight Fi: 4. Effect of drying and heating of a different (from Fig. 3) extract upon its elution pattern HÍATE0 T0 K°C its m" Molecular Weight L U Fig. 3. Effect of drying and heating of an extract upon elution pattern HOT MNO - /U HEÁTE0 TO WC HEATED T0 SO'C
