During the start-up period, 2 L of mixed sludge and 2 L of synthetic textile wastewater were added into the reactor system giving the working volume of 4 L with 5.5 g/L of sludge concentration after inoculation. The system was supplied with external carbon sources consisting of glucose, sodium acetate and ethanol with substrate loading rate of 2.4 kg COD/m3-d. The operation of the system started with 5 min filling of wastewater entering from the bottom of the reactor. The operation then continued with the react phase followed by 5 min settling, 5 min decanting and 5 min of idle time. The react time varies depending on the hydraulic retention time set for the system. Figure 3 shows the steps involved in one complete cycle of the hybrid biogranular system. During the biogranules development, the HRT of the reactor was set for 6 hours for one complete cycle. This will give a react time of 340 minutes. The react phase is divided into equal anaerobic and aerobic react periods. Table 2 shows the successive phase for one complete cycle of the reactor system.

Reaction Phase
Anaerobic Aerobic

The operation of the reactor system was designed with intermittent anaerobic and aerobic react phases. The reaction phase started with an anaerobic phase followed by an aerobic phase. The reaction phase was repeated twice. During the anaerobic react phase, the wastewater was allowed to circulate from the upper level of the reactor and returned back through a valve located at the bottom of the system. The circulation process was carried out using a peristaltic pump at a rate of 18 L/h. The circulation system was stopped at the end of the anaerobic phase. The circulation process is required to achieve a homogeneous

distribution of substrate as well as a uniform distribution of the granular biomass and restricts the concentration gradient. The DO concentrations remained low during the anaerobic condition (0.2 mg/L) and reached saturation during the aerobic phase. The superficial air velocity during the aerobic phase was 1.6 cm/s. The system was operated without pH control causing variation in the range of 6.0 to 7.8 during the react phase.

In order to observe the changes on the characteristics of the biogranules due to the variations of HRT during textile wastewater treatment, the development of biogranules with sizes in the range of 0.3-2.5 mm was inoculated into the bioreactor at a ratio of 1:4 of the working volume of the reactor system. 1 L of acclimated mixed sludge was also added into the reactor system. The MLSS and MLVSS concentrations during the start-up of the experiment were 23.2 g/L and 18.4 g/L respectively. The operation steps of one complete cycle of the reactor system are shown in Table 3.

The morphological and structural observations of the granules were carried out using a stereo microscope equipped with digital image management and analyzer (PAX-ITv6, ARC PAX — CAM). The microbial compositions of the biogranules were observed qualitatively with a scanning electronic microscope (FESEM-Zeiss Supra 35 VPFESEM). The biogranules were left dried at room temperature prior to gold sputter coating (Bio Rad Polaron Division SEM Coating System) with coating current of 20 mM for 45 s. The microbial activity of the biogranules was

determined by measuring the oxygen utilization rate (OUR) following Standard Methods (APHA, 2005). The physical characteristics of the biogranules including settling velocity, sludge volume index, granular strength were measured throughout the experiment.

An initial value of 15 mL influent sample was taken from the influent tank before a new cycle operation started, while another 15 mL of the effluent sample was taken from the effluent tank after the effluent was released during the decanting phase as the final values. Samples were centrifuged for 5 min at 4000 rpm at 40C in order to pellet down all of the suspended particles from the samples. The supernatant was used to measure the removal performance of the COD, color and ammonia removal. All of the measurements for COD, color and ammonia were performed according to Standard Method (APHA, 2005).

10 mL of sample was taken from the top portion of the reactor about 10 minutes after the filling stage ended and 10 mL of sample was taken from the effluent after the decanting stage for the measurement of the suspended particles in the influent and effluent. Another 10 mL of sample volume was taken during the aeration phase for the analysis of MLSS and MLVSS, which were measured according to Standard Methods (APHA, 2005).

The bed height of the biomass in the reactor was measured twice a week in order to estimate the SVI. The bed height was determined immediately after the settling time ended and before the wastewater was drained out during the decanting time. The SVI value can be calculated by measuring the bed volume of the sludge biomass in the reactor divided with the dry weight of the biomass in the reactor. The bed volume is the bed height of the sludge biomass that settled in the reactor 5 minutes after the aeration phase stopped. The bed volume is obtained by multiplying the bed height with the surface area of the bed column. The measurement of the SVI and the sludge retention time were calculated according to Beun et al. (1999). The settling velocity was determined by recording the average time taken for an individual granule to settle at a certain height in a glass column filled with tap water (Linlin et al., 2005).

Determination of the biogranules’ strength was based on Ghangrekar et al. (1996). Shear force on the biogranules was introduced through agitation using an orbital shaker at 200 rpm for 5 minutes. At a certain degree of the shear force, parts of the biogranules that are not strongly attached within the biogranules will detach. The ruptured biogranules were separated by allowing the fractions to settle for 1 minute in a 150 ml measuring cylinder. The dry weight of the settled biogranules and the residual biogranules in the supernatant were measured. The ratio of the solids in the supernatant to the total weight of the biogranular sludge used for biogranular strength measurement was expressed as the integrity coefficient (IC) in percent. This percentage indirectly represents the strength of the biogranules. The higher the IC values the lesser the strength of the biogranules and vice versa.