Toxicity of Mount St. Helens ash leachate to a blue-green alga

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TABLES TOXICITY OF MOUNT ST. HELENS ASH LEACHATE TO A BLUE-GREEN ALGA

The May 18, 1980, eruption of Mount St. Helens volcano in Southwestern Washington covered large areas of Washington, Idaho, and Montana with volcanic ash with accumulation of more than 50 mm in parts of central Washington (fig. 1). To ascertain the possible effects of the ash on water quality, ash samples were collected from several locations for chemical analysis of their distilled water leachates. The results indicate that, relative to natural sur:face waters in tbe affected area, the leachates contain high concentrations of several potentially toxic trace elements, specifically manganese (Mn), zinc (Zn), copper (Cu), and cadmium (Cd) (table 1).

In addition, the leachates contain large concentrations of important algal nutrients (phosphate, nitrate, ammonia) and as much as 200 milligrams of dissolved organic carbon per liter (mgC/L) (Taylor and Lichte, 1980). Chemical analyses of the organic compounds in volcanic ash collected at Richland, Washington, indicate that many of the identified components (for example, dicardoxylic acids, n-alkanes, 4-methaxybenzaldehyde, and trimethylphenanthrene) could have originated as breakdown products of pyrolized plant material (Pereira and others, 1980). These organic products may have formed during the eruption as a lateral blast of superheated gases and hot ash flattened about 40 square kilometers of coniferous forest. J. A. Leenheer (oral communication, 1980) of the U.S. Geological Survey reported that the concentration of organic compounds in leachate from ash varied with location, with higher concentrations near the periphery of the ash fall area than toward its center. In this context, Richland, Wash., is on the southern edge of the ash fall about 140 miles east of Mount St. Helens (fig. 1).

Because of the large concentrations of trace elements and dissolved organic compounds in the leachates, concern arose over the possible effects of the ash on aquatic ecosystems, particularly in the many lakes of the area. Therefore, a preliminary bioassay experiment was conducted using volcanic ash leached with a synthetic solution simulating natural lake water. The organism selected for the test was Anabaena jlos-aquae, a common blue-green algae known to be sensitive to trace metals (McKnight and Morel, 1980). Results showed that leachate from volcanic ash collected in

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the Richland, Wash., area was lethal to Anabaena flos-aquae at dilutions of up to 1~100. Even at weaker dilutions of up to 1:500, the ash leachate caused abnormal morphologies in the algal cells. These biological effects are particulary startling, because the noted laboratory dilutions approximate natural levels of chemical constituents that could be leached from volcanic ash; for example, a 1:250 dilution corresponds to one centimeter of ash falling into a lake with a mixing depth of 2.5 meters.

The objectives of the present study are to: (1) determine whether trace elements, organic compounds, or both, are the toxic components in the leachate from volcanic ash deposited at Richland; (2) isolate and identify the trace metals or organic compounds causing the toxicity; and (3) determine if leachate from volcanic ash deposited near Moses Lake, Wash., is also toxic to blue-green algae. The Moses Lake region in central Washington has many lakes and received one of the thickest ash deposits (fig. 1). The hypothesis that the observed areal variations in organic and inorganic composition of the ash (Leenheer, oral communication, 1980) would result in corresponding variations in toxicity of leachate from volcanic ash was also tested.

To aid the reader, a glossary of technical terms is included at the end of this report.

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To assess the effects of the Mount St. Helens ash on blue-green algal populations in lakes, bioassay experiments were conducted with Anabaena flos-aquae. The general approach in bioassay work is to study the interactions between organisms and their environment under known, controlled laboratory conditions where results can be more readily interpreted than under complex field conditions. The controlled conditions in the present study consisted of constant light and temperature, and a culture medium prepared from laboratory chemicals to simulate the natural aquatic environment. Several simultaneous bioassay experiments were conducted using different chemical fractions of the leachate from volcanic ash to determine the chemical characteristics of the toxic constituents. The flow chart in figure 2 shows the experimental design used in the experiments. Essentially, the experiment consisted of: (1) testing the toxicity of the unfractionated leachate; (2) passing the leachate through a cation-exchange resin;, and (3) testing the toxicity of the effluent from which most trace metals had been removed; and (4) testing the toxicity of the trace metals removed from the leachate by the cation-exchange resin. The specific methodologies are described below in the following

sequence: (1) culture of blue-green algae; (2) leaching of volcanic ash samples; (3) fractionation of leachates; and (4) addition of leachate fractions to algal cultures.

Anabaena .flos-aquae UTEX 1444 was grown in 250 milliliters (ml) of synthetic freshwater culture medium (WC medium) in 500 ml polycarbonate Erlenmeyer flasks at 23°C under continuous light. Synthetic culture medium provides more optimal conditions than most natural waters, permitting Anabaena to grow faster, and providing for more reproducible and controlled experiments. Other reasons for using synthetic culture medium are the large variations in chemistry of lake waters in the study area (table 1) and the difficulty in transporting and preserving natural waters for laboratory study. Growth was monitored by measl1rernent of

TIME

F5 the concentration of chlorophyll a in the cultures every other day and by microscopic examination of the cultures.

Increase of biomass in laboratory cultures of micro-organisms normally follows a sequence of slow growth (lag phase), rapid growth (exponential growth phase), and then no growth (stationary phase) (fig. 3). Toxicants may affect micro-organisms in any growth phase; for example, the lag phase may increase in length as cells adapt to the stress, the growth rate during the exponential growth phase may decrease, or the maximum biomass reached during the stationary phase may be lowered, owing to inhibition of nutrient assimilation.

In the experiment with volcanic ash collected at Richland, leachate was eluted with WC medium from a column packed with 80 grams of ash and

PHASE

saturated for 14 hours at 23oe with 100 ml of we medium (without trace metals orfulvic acid added). In the experiment with volcanic ash collected at Moses Lake, 172 grams of ash were leached with 150 ml of We medium for 12 hours at 23°e and the leachate was eluted from the column with we medium. The amount of ash used in each experiment was governed by the amount of ash available for study. A Millipore HA 0.45~-tm filter, previously leached in we medium, was placed at the bottom of the column to retain particulate material. The leachates were stored in glass containers at 4°e.

In table 1, the dissolved chemical constituents of unaltered we medium and leachates from volcanic ash collected at Richland and Moses Lake are compared with those of four lakes in the ash fall area. The amounts of trace metals and organic material leached expressed as micrograms per gram of ash, are also shown. The analysis for major cations and trace metals was done by inductively coupled plasma emission spectroscopy and the dissolved organic carbon was determined with a Beckman 915 carbon analyzer. Several of the measured constituents (Mn, eu, Zn, ed, and organic carbon) were present in concentrations high enough to be possibly toxic to algae (see Hutchinson, 1957; Bartlett and others, 1974). Both leachates have high concentrations of eu, ed, and Zn. Almost twice as much organic carbon was leached per gram of ash from the Richland sample, which is in accord with the trend reported by Leenheer (see page Fl) toward' highest organic contents in samples from the periphery of the ash fall area.

In order to remove cationic trace metals and organic compounds, both the volcanic ash from leachates collected at Richland and Moses Lake were passed through a 30 ml column of sodiumsaturated cation-exchange resin previously extracted with methanol. Owing to the different amounts of leachate available from the previous step, different volumes were fractionated for the Richland and Moses Lake samples.

A 25 ml sample of the leachate from the ash collected at Richland was applied to the column and 50 ml of leachate and distilled water was eluted with distilled water. Only 67 percent of the organic

F6 carbon in the leachate was recovered in the 50 ml. Another 50 ml of distilled water was passed through the column bringing the total recovery of dissolved organic carbon to 83 percent. These two 50 ml effluent fractions were combined resulting in a fourfold dilution of the leachate. As shown in table 2, most of the trace metals were removed by the cation-exchange resin.

A 50 ml sample of the leachate from ash collected at Moses Lake was applied to an identical column and eluted with distilled water and 100 ml of column effluent was collected. The recovery of dissolved organic carbon in the effluent was 93 percent, much higher than with the Richland leachate (table 2). Most of the trace metals in the Moses Lake leachate were retained by the cation-exchange resin.

The leachate fractions were filter sterilized and added to duplicate flasks of culture medium before inoculation with Anabaenaflos-aquae. Because the ash leachate from ash collected at Moses Lake had a lower concentration of dissolved organic carbon, the amounts of leachate added were scaled to approximate the concentrations of dissolved organic carbon in leachate from ash collected at Richland that were toxic in the preliminary experiment. In the two main experiments, a concentration of 2 milligrams dissolved organic carbon per liter (mg e/L) was used to correspond to the 1:50 dilution that was completely inhibitory, and a concentration of 0.2 mg e/L was used to correspond to the 1:500 dilution that caused abnormal cell morphologies, but did not inhibit growth.

Table 3 presents the concentrations of trace metals and dissolved organic carbon obtained in the culture media by adding different volumes of unmodified leachate from volcanic ash, cation-exchanged leachate, and synthetic trace metals. The synthetic trace-metal solutions were formulated to reproduce concentrations of trace metals in the leachates which might be toxic to algae at a 1:100 dilution. Manganese was highest in concentration among the trace metals in both leachates. However, even at the 1:25 dilution, the concentrations of manganese approximated those in natural waters of the affected area (compare table 3 to table 1).

Figure 4 shows the growth curves for the experiment with leachate from volcanic ash collected at Richland. The graphs show similar growth rates for the control, cation-exchanged leachate, and synthetic trace metal cultures (0.62, 0.64, and 0.58 day- , table 4). As in the preliminary experiment, the unfractionated leachate had a lethal effect on Anabaena flos-aquae at a concentration of 2 mg C/L of dissolved organic carbon (see table 3). The cultures exposed to the unfractionated leachate at a concentration of 0.27 mgC/L had a 7-day period of no net growth (lag phase), prior to an exponential growth phase with replicate growth rates of 0.51 and 0.69 day- • In comparison, a 2-day lag phase had been observed at a concentration of 0.20 mgC/L of leachate (1:500 dilution) in the preliminary experiment.

The results of this experiment corresponds to the third possibility depicted in figure 2, wherein the leachate is toxic, but the cation-exchanged leachate and the synthetic trace metals are not. The most likely conclusion is that the toxicants are part of the 17 percent of organic carbon that was retained by the cation-exchange column. Several types of organic compounds could be retained: (1) cationic-organic compounds such as amines; (2)

F7 strongly hydrophobic (sparingly soluble) organic compounds such as phenols that interact with and are retained by the resin; or (3) metal-organic complexes that are retained by interaction of the metal with the cation-exchange resin. Further experiments are planned to distinguish between these possible types of organic compounds and to obtain a sufficiently large sample of toxicant for more detailed characterization.

Photomicrographs (fig. 5) show Anabaena flos-aquae filaments from the experiments with leachate from volcanic ash collected at Richland. Filaments from the cultures with cation-exchanged leachate and synthetic trace metals were similar in appearance to those in the control cultures. In the cultures with the higher concentration of unfractionated leachate, where no growth occurred, only

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FlO occurs in filamentous blue-green algae- were observed in all cultures, except those exposed to the unfractionated leachate from volcanic ash. Perhaps the toxicants act at the cell wall and interfere with nitrogen assimilation.

Duplicate cultures of fresh, untreated WC medium were inoculated with abnormal Anabaena flos-aquae cells from the Richland experimental cultures. The cultures inoculated with Anabaena flos-aquae cells initially exposed to the lower concentration of leachate grew within a week, whereas the cultures innoculated with cells exposed to the higher leachate concentration grew after three weeks. No abnormal cells were observed in the final cultures, indicating that the effect of the toxicants on the morphology of the blue-green algal cells is reversible.

Figure 6 presents the growth curves from the experiment in which different fractions of leachate from volcanic ash collected at Moses Lake were added toAnabaenaflos-aquae cultures. None of the leachate additions had significant effects on growth or final biomass, nor caused changes in cell morphology. The data in table 4 shows no significant difference in growth rate computed from day 2 to day 4.4 among the various cultures.

The failure to observe any toxic effect of the leachate from volcanic ash collected at Moses Lake is not surprising given that (1) the toxicants in the leachate from volcanic ash collected at Richland were apparently part of the 17 percent of the dissolved organic carbon retained by the cation-exchange column and (2) this fraction contributed only 7 percent of the total dissolved organic carbon to the leachate from volcanic ash collected at Moses Lake. The toxicants may not be present in the leachate or they may occur in relatively much lower concentrations.

The data reported here confirm that leachate from volcanic ash collected at Richland is toxic to Anabaenaflos-aquae at large dilutions. In addition, the results indicate the toxic substances are probably organic compounds retained on a cation-exchange resin, rather than uncomplexed trace metals such as Mn, Zn, Cu, and Cd; or anionic or hydrophillic (readily soluble) organic compounds. The toxic constituents retained on the cation-exchange resin could be one or more of the following types: (1) cationic-organic compounds such as amines; (2) strongly hydrophobic (sparingly soluble) organic compounds such as phenols that interact with and are retained by the resin; or (3) metal-organic complexes which are retained by interaction of the metal with the cation-exchange resin. Further experiments are being conducted to provide more specific identification of the toxic compounds.

Results indicate that the leachate from ash collected at Moses Lake has neither an inhibitory nor stimulatory effect on the growth of Anabaenaflos-aquae. This finding is consistent with the contrasting results from the experiment using leachate from volcanic ash collected at Richland, because the fraction of that leachate shown to be toxic occurs at a much lower concentration in the Moses Lake sample. The toxicants found in the leachate from the volcanic ash collected at Richland may either be absent or occur in much lower, nontoxic concentrations in the leachate from ash collected at Moses Lake.

Visual observation and other data (Fruchter and others, 1980) have shown that the volcanic ash was fractionated (sorted by weight of particles) as it was carried eastward from Mount St. Helens. The lighter materials were carried farthest, and this distribution is reflected in data showing highest concentrations of dissolved organic carbon in

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samples from the periphery of the ash fall area (Taylor and Lichte, 1980; Pereira and others, 1980). Delineation of the geographic distribution of the toxicants is planned for future study. This information will help identify lakes and streams most likely to be affected by volcanic ash deposition. Inh~bition of blue-green algal growth in Liberty Lake, Wash. (see fig. 1), which has a history of nuisance blue-green algal blooms, has already been reported (W. H. Funk, Washington State University, written communication, 1980).

The failure to observe toxicity in laboratory experiments with leachate from volcanic ash collected at Moses Lake is consistent with field observations of algal populations in several lakes from the area. Soap Lake and Lake Lenore, northwest of the town of Moses Lake, have been studied for several decades and their summer phytoplankton assemblages during 1980 were similar to those in previous years (Walker, 1975; W. T. Edmondson, University of Washington, oral communication, 1980). Moses Lake, itself, had the usual dense, blue-green algal bloom. In Warden Lake, south of the town of Moses Lake, the 1980 phytoplankton assemblage was dominated by diatoms and chysophytes, the same as reported (Dione and others, 1980) for previous years.

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