Agricultural fertiliser from brewery effluent – the recovery of nutrients from the biomass of activated sludge and high rate algal pond treatment systems

The disposal of waste biomass generated from biological wastewater treatment plants is a costly process and poses environmental threats to the receiving environment. This study aimed to determine the suitability of algae and waste activated sludge (WAS) produced from a brewery effluent treatment system as a fertiliser in agriculture. The change in soil characteristics and the growth of a crop fertilised with algae or WAS was compared with a conventional inorganic fertiliser. Swiss chard plants (Beta vulgaris) fertilised with anaerobically digested (AD) algae or WAS had a significantly higher mean biweekly yield (5.08± 0.73 kg/m) when compared with the inorganic fertiliser control (3.45± 0.89 kg/m; p< 0.0001). No difference was observed in the soil’s physical fertility when algae or WAS were applied to the soil (p> 0.05). The nitrogen applied to the soil from algae and WAS biomass appeared to leach out of the soil less than the nitrogen supplied by inorganic fertilisers. The application of WAS or algae on soil increased the soil’s sodium concentration and sodium absorption ratio from 774.80± 13.66 mg/kg to 952.17± 34.89 mg/kg and 2.91± 0.04 to 3.53± 0.13, respectively. Regulations on the application of algae or WAS on agricultural soils should be altered to consider the limit values for sodium.


INTRODUCTION
Organic and inorganic pollutants are converted into biomass during the treatment of wastewater in biological systems such as high rate algal ponds (HRAP) and activated sludge (AS; Tchobanoglous et al. ). This biomass (biosolids) is separated from the treated effluent and disposed of. The disposal of waste biomass is a costly process and poses environmental threats if it is not carried out correctly (Hospido et al. ). Tchobanoglous et al. () state that the 'management of the solids and concentrated contaminants removed by effluent treatment has been, and continues to be, one of the most difficult and expensive problems in the field of wastewater engineering'. Practices and technologies need to be developed that allow the sustainable disposal of biomass, especially where the recovery or reuse of resources trapped in the biomass are considered (Singh & Agrawal ). Wastewater biosolids are nutrient-rich organic solids which have the potential to be used for fertiliser, soil amendment or energy production (Dobhal & Singhal ; Elalami et al. ).
Biosolids can supplement or replace commercial fertilisers as they contain nitrogen, phosphorus and various micro-nutrients such as copper, zinc, molybdenum, boron, calcium, iron, magnesium, and manganese (Singh & Agrawal ; Engida et al. ). The organic matter content can be used to increase the cation exchange capacity (CEC) of the soil and can aid in amending soils that are carbon-depleted (Engida et al. ), and thus can potentially improve crop production. For example, the yield of maize, amaranthus, cowpea and crossandra fertilised with sewage sludge was the same as, or higher than, that of plants fertilised with a commercial inorganic fertiliser (Chitdeshwari et al. ; Engida et al. ). The yield of barley fertilised with waste activated sludge (WAS) applied to soils at 15 t/ha dry weight was significantly higher than barley fertilised with a commercial inorganic fertiliser (Antolín et al. ). After four years of annual application of WAS from a sewage treatment facility, the soil's CEC, total organic carbon and available nitrogen increased significantly; however, so did the heavy metal concentration in the harvested grain (Antolín et al. ).
The majority of studies have reported an improvement in physical properties when WAS is applied to soils (Epstein ; Tsadilas et al. ; Lu et al. ). The application of WAS to soils increases its organic matter content, which aids in stabilising the soil structure by increasing interparticulate cohesion within aggregates and enhancing their hydrophobicity (Diacono & Montemurro ). Municipal solid waste applied to the soil, every two years, increased soil aggregate stability by 29%, thus increasing its resistance to erosion (Annabi et al. ). The application of sewage sludge to the soil has been shown to improve its water holding capacity, porosity and the bulk density of the soil (Epstein ; Ojeda et al. ).
Anaerobic digestion is a recommended stabilisation step prior to the land disposal of waste sludge as it allows the recovery of carbon into an energy source (biogas) and can increase the availability of nutrients to plants (Tchobanoglous et al. ). This process results in the conversion of protein-bound nitrogen to ammonia which can be utilised by plants and can increase the nutrient availability of biosolids. Anaerobically digested WAS had higher phosphorus availability than heat-dried WAS (Lyberatos et al. ). To date, there is only one publication which compares the use of AD to increase the nutrient availability of WAS to plants (Warman & Termeer ). Sludge pre-treated using AD resulted in a higher yield of Zea mays when compared with sludge that was composted (Warman & Termeer ). No other current literature has compared the use of AD to possibly increase the fertiliser value of effluent-grown algae and to document the subsequent effect on soil fertility.
High rate algal ponds and AS are effective brewery effluent treatment technologies which both produce a biomass than can be used in agriculture. The suitability of these biomasses as a fertiliser needs to be assessed and can aid in deciding which technology is favourable in a particular situation. Algae have resilient cell walls which can decrease their decomposition rate and thus reduce the availability of nutrients to plants when compared with WAS (Markou et al. ). Algal biomass cultured in piggery wastewater significantly increased the growth of wheat when compared with unfertilised treatments ( Jenkins et al. ), however this was a pot experiment and only carried out for four weeks. This is the first study which compares the use of algae and WAS as a fertiliser replacement, where both the algae and the WAS are produced from effluent treatment systems that have been used to treat the same effluent.

Aims and objectives
The aim of this study was to determine the suitability of algae and WAS produced from a brewery effluent treatment system as a fertiliser in agriculture. This was done by comparing the change in soil characteristics and the growth of a crop fertilised with algae or WAS with a conventional inorganic fertiliser. This study will also determine what effect an AD pre-treatment step may have on the fertiliser quality and the fertility of the receiving soil.
The objectives of this study were to: • compare plant growth and soil characteristics of plots fertilised with WAS, algae or inorganic fertiliser; • compare plant growth and soil characteristics between soils fertilised with anaerobically digested and non-anaerobically digested algae or WAS; and • determine the suitability of anaerobically digested and non-anaerobically digested WAS or algal biomass as an inorganic fertiliser replacement.

MATERIALS AND METHODS
Untreated brewery effluent was screened through a 500 μm drum filter (Autrex Industrial Screening, serial no. A 140/02, model no. R 015) to remove solids and then anaerobically digested in an up-flow anaerobic sludge blanket reactor.
After AD the effluent was polished in a commercial AS system or a pilot HRAP system.
The majority of the post-AD effluent (800 m 3 /d) was treated in an AS system consisting of a rectangular aeration basin and clarifier operated by an independent commercial company (Proxa Pty Ltd, South Africa). The aeration basin was operated at a hydraulic retention time (HRT) between 0.4 and 0.5 days, while the dissolved oxygen concentration was maintained around 0.8 mg/l. Effluent was decanted from the aeration basin into a clarifier operated at an HRT between 0.3 and 0.4 days, which separated the suspended solids from the treated effluent. Settled sludge from the clarifier was also disposed of into the municipal sewage works, and this process was called 'sludge wasting'. This was done to maintain the desired concentration of microorganisms in the aeration basin of 350-400 ml/l volume of settleables. Sludge wasting was performed once or twice a week depending on the volume of settleables in the aeration basin, and was done when the volume of settleables increased above 450 ml/l and was stopped when it decreased to 300-350 ml/l. Effluent entering the HRAP had undergone AD and stabilisation in a primary facultative pond (PFP) operated at a HRT of four days. Effluent from the PFP was decanted into two identical parallel HRAP systems, consisting of two ponds in series. The first pond of each system was 25 cm deep, with a surface area of 14.8 m 2 and a volume of 3,700 l. Effluent decanted from the first pond to the second pond, which was 11.5 cm deep, with a surface area of 15 m 2 and a volume of 1,700 l. Each pond had a stainless-steel paddle wheel which continuously stirred the effluent and kept the algal cells in suspension. Paddle wheels moved HRAP effluent at an approximate velocity of 4.15 m/s and 6.10 m/s in ponds 1 and 2 respectively. Both algal systems were fed 1,800 l/d of effluent, which equated to an HRT of three days. The HRAPs were fed continuously during daylight hours (08:00-17:00), while feed stopped during the remaining 15 h. Post-HRAP effluent, from both systems, flowed into two algal settling cones which separated the algal biomass from the treated effluent.
The settled algae and WAS slurries were used directly as a fertiliser or anaerobically digested and then used as a fertiliser in the crop production trial. to the soil at a nitrogen application rate equivalent to 80 kg/ha (Laboski & Peters ), two days before the soil was planted, and they were mixed into the top five centimetres of the soil.

TREATMENTS
Two semi-continuously-fed anaerobic digesters were used to digest the settled WAS and algal biomass. Both were made of plastic drums with a total volume of 220 l, a head space of 60 l and an operating volume of 160 l. They were stirred with a submersible mixer (Sobo, WP 400M, South Africa) for five minutes every half an hour, and were situated in a temperature-controlled room (37 C).
The digesters were seeded with sludge obtained from a biogas-producing up-flow anaerobic sludge blanket reactor at Ibhayi Brewery. They were fed once a day following the removal of the equivalent volume of digestate. The settled algae and WAS fed to the digester had a total solids content of 25 g/l. Each digester was fed 9.5 l of either algae or WAS per day resulting in a feeding rate of 1.5-2.0 g of TS/l reactor /d and an HRT of 16 d. These two digesters were operated for 60 days, before the digestate was collected and applied to the soil as a fertiliser. were filled with the amended soils. Each soil amendment treatment was randomly assigned to three raised beds, such that the treatments were replicated three times with a replicate consisting of a single raised bed.

EXPERIMENTAL SPECIES, SYSTEM AND IRRIGATION
They were irrigated once a day, except during rain or directly thereafter (Laboski & Peters ), and received a total of 377 mm of water over the 13-week growth trial (275 mm irrigation and 102 mm of rain). Plants were irrigated via a drip irrigation system receiving a maximum of 5 mm of water per irrigation to minimise leaching (Laboski & Peters ).

DATA COLLECTION
A sample of the un-digested and anaerobically digested WAS and algae that were applied to the soil were subject to elemental analysis (inductively coupled plasma massspectrometry and X-ray fluorescence) at an independent laboratory (Central Analytical Facilities, Stellenbosch University, South Africa).
At the beginning of the trial, ten plants were randomly taken from the population of seedlings that were used for the experiment, and were weighed to determine the mean wet starting mass (0.1 g). After five weeks, the plants were ready for harvesting, when all the large, fully expanded leaves from each plant were removed and the wet weight weighed (0.1 g). This was repeated every two weeks until the experiment was terminated, after 13 weeks. At the end of the trial, all the above-ground biomass was harvested and wet weight weighed (0.1 g). where the holes were unblocked. The vessel was left to drain for 30 min and the amount of water collected was measured. Air-filled porosity was calculated using Equation (2) (ISO ). Directly after the AFP test the vessel was placed in a drying oven at 105 o C and allowed to dry for a minimum of 24 h, until a constant mass was achieved.
Water holding capacity was calculated using Equation (3) (ISO ), and bulk density was then calculated using Equation (4) (ISO ).
Air filled porosity (%) ¼ (volume drained/volume of soil) × 100 ( 2) Water holding capacity (%) Bulk density (g=cm 3 ) ¼ dry weight=volume (4) Soil aggregate stability was measured, in each replicate, at the beginning and end of the experiment using five grams of 2-5 mm aggregates (Le Bissonnais ). Samples were placed in distilled water and allowed to stand for ten minutes. The distilled water was then removed with a pipette and the aggregates transferred onto a 0.05 mm sieve which was immersed in ethanol and shaken five times with a gentle regular helical rotation movement. The where d was the mean diameter between the two sieves (mm) and m was the weight fraction of aggregates remaining on the sieve (%).
The chemical analysis of the soil was determined at the start of the trial (n ¼ 5 The sodium adsorption ratio (SAR) of the soil was calculated using Equation (6), where sodium, calcium, and magnesium are expressed in milliequivalents per litre (meq/l), obtained from a saturated paste soil extract (Qadir et al. ): Samples of anaerobically digested and un-digested WAS or algae applied to the soil were tested for Escherichia coli.
Similar analyses were repeated on soil and leaf samples taken from each replicate every four weeks at the Ibhayi Brewery laboratories (IS 17994 method; ISO ).

STATISTICAL ANALYSIS
The experimental design allowed for: (1) a multifactor analysis of variance (ANOVA) where the treatments included two soil-amendments (algae and WAS), both of which were either subject to AD or left un-digested; and (2) a one-way ANOVA that included a comparison of the four treatments described above and a fifth inorganic-fertiliser treatment that acted as the reference-control. If no significant interactions were observed between soil-amendment factor and/or AD pretreatment factor (multifactor ANOVA), then the statistical analysis generated from the one-way ANOVA/Kruskal-Wallis ANOVA was used. All analyses were carried out at p < 0.05 and, when differences were found, a Tukey's multiple range analysis was used at p < 0.05. Data collected over the course of the trial were compared using multifactor repeated measures or one-way ANOVA or a non-para-

RESULTS
WAS and algal biomass applied to the soil were within the heavy metal limit for application to agricultural land (   The physical properties of the soil were not influenced by an interaction between factors of biomass source and AD pre-treatment (multifactor ANOVA, p > 0.05). After 13 weeks, there was also no significant difference in the bulk density, porosity, water holding capacity, infiltration rate and mean weight diameter of soils subject to the five  Anaerobically digested algae (AD-algae), anaerobically digested WAS (AD-WAS).
Superscripts (a, b) in the same row represent significantly different treatment means (one-way ANOVA/Kruskal-Wallis, p < 0.05).
All measured soil chemical concentrations were not influenced by an interaction between biomass type and AD pre-treatment (multifactor ANOVA, p > 0.05). The zinc concentration of the soil was significantly higher in the algae-fertilised soils when compared with WAS-fertilised soils (  Anaerobically digested algae (AD-algae), anaerobically digested WAS (AD-WAS).  When applying sludge to soils the possibility of contaminating them with heavy metals exists. Of the heavy metals analysed, zinc was the only element that was higher in  Anaerobically digested algae (AD-algae), anaerobically digested WAS (AD-WAS).
Superscripts (a, b) in the same row represent significantly different treatment means (multifactor ANOVA, p < 0.05).
application of sludge from WWTPs should not have any detrimental effect on the soil's chemical fertility (Zerzghi et al. ).
Sodium contamination of agricultural soils is the leading cause of rendering soils unsuitable for agriculture (Qadir et al. ; Muyen et al. ). In this study, the sodium concentration and SAR of soils fertilised with algae or WAS were significantly higher than those of the soils fertilised with inor-  Anaerobic digestion is able to recover a portion of the carbon in waste biomass into a fuel source (methane) before it is applied to the soil. The carbon concentration of soils fertilised with AD sludge or algae was significantly lower than soils fertilised with undigested sludge or algae.
During AD the carbon content of organic matter is converted into methane and carbon dioxide, thus decreasing its carbon concentration (Tchobanoglous et al. ).
Anaerobic digestion is a recommended pre-treatment step

CONCLUSION
These waste products can be utilised as an inorganic fertiliser replacement when applied to the soil at the same nitrogen loading rate and can even increase crop yield.
The nitrogen applied to the soil from algal and WAS biomass appeared to leach out of the soil less than the nitrogen (ammonia and nitrate) supplied by inorganic fertilisers. These biomasses offer a good alternative to inorganic fertilisers as the slow nutrient release will aid in reducing the nutrient contamination conventional inorganic fertiliser-based agriculture has on surrounding water bodies.
Anaerobic digestion is a viable pre-treatment step for sludge and algal biomasses as it recovers a portion of the carbon as an energy source and does not decrease their quality as fertilisers. The application of algae generated from a brewery effluent treatment system increased the soil sodium concentration, so it is recommended that regulations on the application rate of sludge to agricultural land should consider the limit values for sodium to avoid its accumulation in the soil and deterioration in soil fertility.