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The effect of salinity on pH mediated abiotic nutrient removal was determined. Nutrient removal occurred at elevated pH in all dilutions of seawater (1:0, 1:1 and 0:1) and in the 1:1 diluted wastewater (Fig, 4.10). Ammonium removal began at pH 8.5 and concentrations decreased linearly with increase in pH in all solutions. However, only 23.5 % of the original concentration had been removed at pH 11.2 (Fig. 4.10a). Ortho-phosphate was totally removed from all solutions, but the pH range over which this occurred depended upon the salinity of the medium. For distilled water the range was between pH 8.7 and pH 10.1 whereas in seawater and the 1:1 dilutions of seawater and wastewater the range was slightly higher, between 9.0 and 10.5 (Fig. 4.10b).

4.4 Discussion

4.4.1 Effect of Temperature on Algal Growth and Nutrient Removal

It was not unexpected that the algal species grown from inocula from the Conviron had temperature optima of 15 ^C, since these species had been screened for nutrient removal and maintained in stock cultures at this tem perature. Following temperature acclimation of cultures on the thermo-gradient, both the temperature optima and ranges of most of the species increased. The optimum of 20 °C is higher than was expected for these temperate species, since the maximum temperature of seawater from St Andrews Bay in the summer was only 15.5 (Chapter 2; Table 2.2). The optimum temperature of algal species in culture are often higher than those in which the species naturally live (Admiraal, 1977). Possibly, the growth of

500 n 400 300- 200- 100- 1--- '--- r 10 11 12 80 n pH

Figure 4.10 Am m onium (a.) and ortho-phosphate (b.) concentrations in distilled water (■-■), 1:1 seawater:distilled

water (♦-♦), seawater (#-#) and 1:1 wastewater:seawater (a-a )

many of these species in St Andrews Bay is temperature limited over much of the year.

Most species grew better at temperatures below their optima than above (Figs. 4.2,4.3,4.4 & 4.5). The inhibitory effect of higher than optimal temperature has been found for many microalgal species (Nelson et al, 1992). During winter months, when seawater temperatures may be as low as 6 °C, temperature may limit the use of some of the best-treating

microalgal species in wastew ater treatm ent system. Tem perature acclimated cultures of three algal species (SA90B2, SA91B33 and SA92B48) removed 1 0 0 % of both ammonium and ortho-phosphate during the two

days culture at 5 °C (Figs. 4.2 & 4.4; Table 4.1), and therefore show promise for use in such a system during the winter.

4.4.2

The different growth responses of 10 of the best-treating algae to a salinity gradient of seawater diluted with M-Q water indicated the variety of salinity tolerances of these marine algae. The similar growth responses of these species on seawater diluted with wastewater are not unexpected because they were selected for their nutrient removal ability from 1:1

seawateriwastewater. The growth of microalgae was probably higher on the wastewater than on the seawater dilutions with E-S nutrients since they use ammonium more readily as a nitrogen source than nitrate (Thompson et al, 1989; Dortch, 1990; Cochlan & Harrison, 1991a, b & c; Raven et al, 1992). The decline in the growth of the endemic strains of Phaeodactylum tricomutum such as SA90B2 at low dilutions of wastewater suggested that for these strains, other properties of the diluted wastewater had more effect on algal growth than salinity. The inhibitory effect of excessive ammonium

cannot be discounted as the ammonium content of undiluted wastewater used in this experiment was 2.4 mol N-NH4+. Algal photosynthesis is

inhibited at ammonium concentrations above 2.0 mol m-3 N-NH3, if the

culture pH exceeds 8.0, when non-toxic NH4+ dissociates to toxic NH3

(Abeliovich, 1980; Azov & Goldman, 1982). The pH in the batch culture of one of the best-treating algae (SA90B2) rose above pH 10.0 after 8 h culture

(Fig. 4.8). However, this would not affect the microalgae in treatment ponds since 1:1 dilution of the wastewater with seawater w ould reduce

ammonium concentrations below this inhibitory level.

Species SA90C3 is typical of many marine microalgae which have a salinity optimum in the range 16-32 %o (Laing & Utting, 1980; Fabregas et al, 1984; Fabregas et al, 1985b). Below the salinity optimum, the decline in growth is proportional to the decrease in salinity (Jiménez et al, 1990). However, other microalgal species have been found to be capable of adapting to changes in salinity, ranging from freshwater 0 %o to oceanic seawater 35 %o (Fabregas et al, 1987a).

4.4.3

The batch culture experiments investigating pH changes during wastewater treatment by one of the best-treating algae have shown that culture pH varies with light:dark cycle (Fig. 4.8). This is a result of bicarbonate removal during high rates of microalgal photosynthesis (Richmond, 1983; Fabregas et al, 1984; Boeder & Hegewald, 1988). In the present study, when the pH of 1:1 diluted wastewater was raised by addition of 1 M NaOH, ortho-phosphate began to precipitate at pH 9.5, and was totally removed at pH 10.5, although more than 70 % of ammonium still remained in solution at pH 11.4 (Fig. 4.8). However, in algal cultures.

over 50 % of ammonium and 70 % of ortho-phosphate were removed before the medium reached pH 9.5 (Fig. 4.8), indicating that nutrient removal in algal batch cultures was mainly due to algal uptake and not due to the abiotic effects of increased pH. Other authors have found ammonium removal from algal cultures to be mainly due to uptake and assimilation (Goldman and Stanley, 1974; M atusiak et al, 1976). In this study ortho-phosphate precipitation was shown to occur at pH 9.5 or above, with concentrations in the dissolved precipitate corresponding to the concentration removed from the culture medium (Fig. 4.9). The salinity of the culture medium also affected the pH at which ortho-phosphate precipitation began. At a higher salinity the pH at which precipitation occurred was increased (Fig. 4.10) which may have been a result of the buffering capacity of seawater (Rebello & Moreira, 1982).

Since the effects of many of the environm ental conditions (temperature, light and nutrients) which influence microalgal growth are interrelated (Vonshak et al, 1982; Henry, 1988; de la Noue & De Pauw, 1988), laboratory studies cannot provide a complete picture of the response of algae to natural conditions (Admiraal, 1977). Therefore, the conclusions reached about growth rate measurements in cultures must be applied with caution to algae growing in the field. Many microalgal species are able to adapt to the conditions under which they are cultured so that their range of tolerance to environmental conditions may be altered with time (Craig et al, 1988). However the experiments described here have shown that salinity, temperature and pH are all conditions that affect the growth of marine algal species and that many of the best-treating microalgal species are capable of growing and removing nutrients over a wide range of temperatures and salinities.