Variation of effluent organic matter (EfOM) during anaerobic/anoxic/oxic (A2O) wastewater treatment processes

Supplementary Material

Gang Tanga,b, Xing Zhenga,*, Xiaolin Lia, Tong Liua, Yan Wangc, Yinliang Maa, Yetong Jia,d, Xiaopeng Qiua, Yi Wanc, Baozhu Pana

aState Key Laboratory of Eco-hydraulics in Northwest Arid Region, Xi’an University of Technology, Xi’an 710048, China

bSchool of Environmental Science and Engineering, Southern University of Science and Technology, Shenzhen 518055, China

cShaanxi Provincial Institute of Microbiology, Xi'an 710043, ChinadKey Laboratory of Integrated Regulation and Resource Development on Shallow Lake of

Ministry of Education, College of Environment, Hohai University, Nanjing 210098, China*Corresponding authorE-mail address: zhengxingde@yahoo.de

PAGE \* MERGEFORMAT1

In section 2.1. Sampling of wastewater and activated sludgeWastewater samples, were collected by composite sampling (mixing three samples from

different sites in the same treatment tank into one sample to reduce the extent of fluctuation caused by different sampling points) on the morning (around 11 AM) of the 24th of every month from a local wastewater treatment plant (WWTP) in Xi’an, China (sketch of the sampling sites is illustrated in S Fig. 1; weather condition of sampling dates is recorded in S Tab. 1).

S Fig. 1. Sketch of A2O processes and wastewater sampling sites

S Tab. 1. Weather condition of Xi’an, China in sampling dates

Sampling dates Highest temperature (°C) Lowest temperature (°C) Weather2018-09-24 25 16 Cloudy2018-10-24 20 12 Cloudy2018-11-24 14 3 Cloudy2018-12-24 4 -1 Overcast2019-01-24 4 0 Cloudy2019-02-24 12 2 Overcast2019-03-24 19 5 Overcast2019-04-24 28 15 Sunny2019-05-24 35 20 Overcast2019-06-24 24 19 Overcast2019-07-24 32 24 Overcast2019-08-24 32 24 Cloudy

PAGE \* MERGEFORMAT1

The WWTP was a separated system which treats only municipal wastewater with a treatment capacity of 200,000 m3/d including removal of organics and nutrients. The effluent quality achieves the Class-1A of Discharge Standard of Pollutants for Municipal Wastewater Treatment Plant (GB 18918-2002) of China. Parameters of the WWTP are as below:

S Tab. 2. Parameters of the WWTP

Parameters ValueHydraulic retention time (HRT) of primary sedimentation tank 1.51 hHRT of anaerobic tank 2.02 hHRT of anoxic tank 5.53 hHRT of oxic tank 8.89 hHRT of secondary sedimentation tank 3.62 hSludge retention time (SRT) of A2O system 15.64 dSRT of oxic tank 8.53 dSludge loading 0.08 kg BOD/kg SS·dMixed liquor suspended solids (MLSS) 3500 mg/LActual oxygen requirement (AOR) 70.15 t/dAir supply 60203.0 m3/h

PAGE \* MERGEFORMAT1

In section 2.2. Determination of conventional water quality indexes

Determination of nutrients, ammonia nitrogen (NH4+-N) and total phosphorus (TP), was

respectively using Nessler’s reagent spectrophotometric method (HJ 535-2009) and ammonium molybdate spectrophotometric method (GB 11893-89), which were recommended by Ministry of Ecological Environment of the People's Republic of China. Specifically, 1.0 mL of potassium sodium tartrate (500.0 g/L, ammonia removed) and 1.0 mL of Nessler’s reagent (10.0 g HgI2, 7.0 g KI and 16.0 g NaOH dissolved in 100 mL deionized water) were mixed well with 50 mL of water samples. After 10 minutes, the absorbance of the mixture was measured by a UV-Vis spectrophotometer (DR6000, HACH, US) at 420 nm wavelength with DI water as background. The concentration of NH4

+-N was determined by the calibration curve obtained by NH4Cl standard solution (R2=0.999). As for TP, 4 mL of K2S2O8 (50.0 g/L) was added to 25 mL of water samples, and the mixture was autoclaved under 121°C for 30min (pressure=1.1 kg/cm2) to digest the TP. Then 1mL of C6H8O6 (100.0 g/L) and 2 mL of ammonium molybdate reagent (130.0 g/L) w ere added to the cooled digested mixture. After 15 minutes, the absorbance of the mixture was measured by a UV-Vis spectrophotometer (DR6000, HACH, US) at 700 nm wavelength with DI water as background. The concentration of TP was determined by the calibration curve obtained by KH2PO4 standard solution (R2=0.999).

PAGE \* MERGEFORMAT1

In section 3.1. Variation of DOC and nutrients along 12 months monitoring period

S Tab. 3. Statistical data of the wastewater temperature in the monitoring year

Months fromSept. 2018-Aug. 2019

Influent Secondary effluentAve Max Min Ave Max Min

Sept. 22.7 24.9 18.7 23.9 26.0 19.9Oct. 20.7 23.0 17.8 21.7 24.1 19.3Nov. 19.2 21.4 17.0 19.9 22.5 18.3Dec. 17.5 19.5 15.9 18.4 19.9 17.1Jan. 16.5 17.2 15.2 17.1 17.8 15.9Feb. 16.5 17.4 15.9 16.9 17.9 16.3Mar. 18.0 18.3 16.7 18.3 18.9 17.0Apr. 19.1 20.1 18.2 20.0 21.4 19.0May 21.6 22.8 19.2 21.5 23.3 19.8Jun. 23.9 24.2 21.3 24.5 25.4 22.6Jul. 24.8 25.4 23.3 25.8 27.6 24.2

Aug. 25.8 26.5 23.9 25.9 27.6 23.0

S Fig. 2. Variation of total phosphorus (TP) in influents, A2O processes and effluents in a year by months (A) and seasons (B)

PAGE \* MERGEFORMAT1

In section 3.2. Variation of molecular weight distribution of EfOM fractions

S Tab. 4. Variation of hydrophobicity of wastewater samples in a year

Seasons fromSept. 2018-Aug. 2019

Hydrophobicity (%)S1 S2 S3 S4 S5

Autumn 15.9 14.9 16.3 18.2 24.2Winter 19.8 18.0 11.6 11.5 14.8Spring 18.5 19.5 15.6 15.6 17.7

Summer 10.1 18.2 11.6 10.8 16.1

PAGE \* MERGEFORMAT1

In section 3.4. Seasonal variation of the microbial community structure

S Tab. 5. Number of observed species in activated sludge from different treatment tanks in different seasons

Seasons fromSept. 2018-Aug. 2019 Treatment tanks Number of observed species

AutumnAnaerobic tank 1770

Anoxic tank 1919Oxic tank 1782

WinterAnaerobic tank 1702

Anoxic tank 1621Oxic tank 1648

SpringAnaerobic tank 1661

Anoxic tank 1754Oxic tank 1744

SummerAnaerobic tank 1654

Anoxic tank 1658Oxic tank 1818

S Fig. 3. Relative abundance of top 10 phyla in A2O processes in different seasons

PAGE \* MERGEFORMAT1

In section 3.6. Synergetic Effect of Temperature and Proteobacteria (SETP)

S Tab. 6. Pearson Correlation Coefficient

Variation of EfOM in Proteobacteria in Environment Water temperatureA1 A2 O A1 A2 O temperature

Variation of EfOM in

A1 1 -0.135 -0.478 -0.325 -0.229 -0.298 -0.439 -0.164A2 -0.135 1 -0.363 0.763 0.592 0.463 -0.460 -0.600O -0.478 -0.363 1 0.306 0.472 0.612 -0.123 -0.233

Proteobacteria inA1 -0.325 0.763 0.306 1 .965 0.924 -0.679 -0.864A2 -0.229 0.592 0.472 0.965 1 0.986 -0.774 -0.920O -0.298 0.463 0.612 0.924 0.986 1 -0.716 -0.869

Environment temperature -0.439 -0.460 -0.123 -0.679 -0.774 -0.716 1 0.956Water temperature -0.164 -0.600 -0.233 -0.864 -0.920 -0.869 0.956 1

A1=anaerobic process, A2=anoxic process, O=oxic process

6