Wastewater not only contains nutrients such as N and P, but also a range of other contaminants that may interfere with microalgal growth. The presence of growth — inhibiting substances probably explains why microalgal growth rates in real wastewaters are often slightly lower than in synthetic wastewaters. Contaminants not only pose a problem because they inhibit microalgal growth, but they can also accumulate in the microalgal biomass and limit the valorization of the microalgal biomass, or fractions thereof. Wastewater can contain a wide range of toxic chemicals such as heavy metals, persistent organic pollutants, and surfactants.

This is particularly the case in industrial wastewaters, although domestic waste­water or animal manure may also contain substantial quantities of pollutants such as heavy metals (Nicholson et al. 1999). Even chemicals with a low toxicity (such as those used in personal care products), may have an inhibitory effect on microalgal growth rates (e. g., Wilson et al. 2003).

Animal manure wastewaters often contain very high concentrations of N present in reduced form as ammonium. Ammonium is converted to free ammonia at high pH and is toxic to many microalgae, with some microalgae inhibited by concen­trations as low as 20 mg L 1 (Azov and Goldman 1982). High concentrations of ammonium may result in ammonia toxicity even when pH is low (Peccia et al. 2013). Some substances do not inhibit microalgal growth, but accumulate in the microalgal biomass and limit valorization. The cell wall of microalgae is often rich in carboxyl, amino, hydroxyl, or sulfate groups. These groups are anionic and can bind metals through ion exchange (Wang and Chen 2009). Microalgae are efficient absorbers of heavy metals and even low concentrations of heavy metals present in wastewater can be absorbed into microalgal biomass. This is not necessarily a problem when the biomass is converted into biofuels using chemical or physical methods. It may be a problem when biological methods are used to convert the biomass into fuel (e. g., anaerobic digestion, fermentation). It may also be a problem when the protein-rich residue of the biomass remaining after extraction of lipids for biodiesel production is to be used as animal feed, as is proposed in the microalgal biorefinery context (Wijffels et al. 2010). Rwehumbiza et al. (2012) showed that metals used for flocculating microalgae remain in the protein-rich residue after extraction of lipids. However, microalgal absorbtion of heavy metals from waste­water can also be an advantage, and numerous studies have demonstrated that microalgae can be applied to remove heavy metals from a variety of wastewaters (see for instance Wilde and Benemann 1993; Gadd 2009). Wastewater may also contain microbial contaminants such as cysts of parasites, infectious bacteria, or viruses. These may also interfere with the use of microalgal biomass fractions as animal feed. However, due to the high pH, high oxygen concentrations, and exposure to light, many harmful microorganisms tend to be inactivated in micro­algal cultures (Davies-Colley et al. 1999).

Many wastewaters of agricultural origin such as piggery waste or anaerobic digestion wastewater often have a dark color. This dark color is predominantly due to the presence of humic substances derived from incomplete breakdown of lignin in plant material (Brezonik and Arnold 2011). This dark color limits the light pene­tration in the water and reduces microalgal productivity (Martin et al. 1985); potentially a 20-30 % lower productivity when compared to growth rates in a non­colored culture medium (De Pauw et al. 1980). In many laboratory studies on production of microalgae in animal manure wastewater, the wastewater is diluted prior to the experiments and inhibition of microalgal productivity by the dark col­oring is barely noticed. However, in large-scale systems, dilution of wastewater with pure water will be unsustainable due to the high water demand. To prevent inhibition of growth by colored substances, the wastewater can be pre-treated with oxidizing agents such as sodium hypochlorite, ozone or hydrogen peroxide, or by coagulants, flocculants, or adsorbents (Markou et al. 2012b; Depraetere et al. 2013). Alterna­tively, nutrients can be separated from the wastewater containing colored substances and then added to the microalgae culture medium. This can be carried out, for instance, using dialysis membranes (Blais et al. 1984). Also nutrients can be sorbed onto zeolites and released from the zeolites in fresh medium (Markou et al. 2014).