Formation of disinfection by-products during the monochloramination of co-existing Microcystis aeruginosa and Cyclops metabolites

Algae have come to the attention of the drinking water industry as a result of the continuing eutrophication of surface water supplies. In China, algal blooms have impacted several major water systems (e.g., Taihu Lake, Chaohu Lake, and Dianchi Lake) in recent years (China Daily 2007). Microcystis aeruginosa is a type of blue-green algae that is distributed all over the world and is present in high quantities and with a high frequency of occurrence in most of the eutrophic water bodies in China. During the period of the outbreak of Microcystis aeruginosa in summer, the monitoring results showed that the number of Microcystis aeruginosa could reach 10⁹count/L (Zhang et al. 2014). Algae and algal metabolites greatly impact the treatment of potable water by causing problems such as poor settling, filter clogging and breakthrough of sand filters by small algae, producing unpleasant tastes and odours, in addition to toxins, and increasing the microbial regrowth potential in distribution systems (Knappe et al. 2004). In addition, algal cells can serve as precursors of disinfection by-products (DBPs) during chlorination. Increased reaction time, chlorine dosage and temperature improved the formation of the relatively stable carbonaceous DBPs (C-DBPs) (e.g., trihalomethane (THM), haloacetic acid (HAA), and chloral hydrate (CH)) and trichloronitromethane (TCNM). The formation of dichloroacetonitrile (DCAN) followed an increasing and then decreasing pattern with prolonged reaction time and increased chlorine dosages (Fang et al. 2010a). Algogenic organic matter, including extracellular organic matter (EOM) and intracellular organic matter (IOM), was rich in organic nitrogen. The MW of organic carbon in EOM and IOM was low. IOM had a higher fraction of free amino acids but lower fractions of aliphatic amines than those of EOM. EOM formed fewer C-DBPs than did IOM. Most trichloro C-DBP yields, such as those of TCM, CH, and 1,1,1-TCP, in chloramination were much lower than those from chlorination. 1,1-DCP yields were higher in chloramination than those in chlorination (Fang et al. 2010b). During the chlorination of Microcystis aeruginosa, the concentrations of THMs and HNMs fluctuated with age. The formation of haloacetonitriles (HANs) during chlorination increased with longer culture ages. The formation of nitrogenous DBPs (N-DBPs), including DCAN and TCNM, did not follow the similar trends during chlorination of algal cells with increasing culture age (Yang et al. 2011).

Zooplankton of the genus Cyclops were observed to propagate excessively in waters that are eutrophiedas, a result of water pollution, especially in reservoirs and fresh lakes that serve as drinking water sources (Sun et al. 2013b). When Cyclops are fully developed, they have a body length of 2.0 millimetres and can be observed by the naked eye causing consumers to have an unsanitary view of the water. In addition, Cyclops may become a disease transmission medium as the host of pathogenic parasites. Cyclops are large in size compared with algae, bacteria, and other microorganisms, which suggests that the Cyclops biomass of amino acids, protein, fat, and other organic matter has greater potential to form DBPs during chlorination (Liu & Fu 2010). Thus, determining how the metabolites produced by these organisms affect water safety and contribute to the production of DBPs is interesting. In addition, Cyclops have strong motility and can easily penetrate filters to enter water supply systems (Cui et al. 2002; Lin et al. 2007). Abundant Cyclops not only pollute the ecological balance but also pose a considerable threat to human health. Many factors have been extensively studied to examine their effects on DBPs formation potential (DBPsFP) during disinfection, including the reaction time, pH, temperature, disinfectant concentration, and precursor properties. The formation of stable THMs and HAAs increased with increasing reaction time and chlorine dosage (Sun et al. 2013b). The formation of DCAN, and TCNM increased and then decreased with increasing chlorine dosages, followed by a continuous decrease with prolonged reaction time (Sun et al. 2013b). 1,1-DCP and 1,1,1-trichloro-2-propanone (1,1,1-TCP) increased with increasing chlorine dosage. 1,1-DCP increased continuously with increasing reaction time, while 1,1,1-TCP decreased with increasing reaction time (Sun et al. 2013b). The pH affected DBP formation differently, with TCM increasing and with DCAA, TCAA, DCAN, and 1,1,1-TCP decreasing (Sun et al. 2013b). TCM, DCAA, and TCAA could accumulate to their respective stable values with a progressive elevation in reaction time and monochloramine concentration. The 1,1,1-TCP content decreased correspondingly with a continuous increase in the reaction time. The formation of CH, TCNM, 1,1,1-TCP, and DCAA initially increased and then decreased with increasing monochloramine doses. A higher temperature resulted in a decrease of CH, DCAN, 1,1-DCP, 1,1,1-TCP, DCAA, and TCAA concentrations. DCAA, TCAA, CH, and 1,1-DCP decreased continuously with increasing pH (Sun et al. 2014).

Chloramine was first used as a disinfectant in water treatment in 1971. Chloramination exceeds the parameters of chlorination in the following three aspects: chloramine has good stability and can sterilize water for long periods, chloramine can slow down the chlorination of humus by free chlorine, and chloramine has a slightly corrosive effect on the pipe network. The monochloramination of chironomid larvae metabolite without generating high levels of most DBPs has been reported (Sun et al. 2013a). TCM is generally considered to be a non-genotoxic carcinogen, whose mechanism of action involves cytotoxicity and regenerative cell proliferation (IARC 1999). TCAA can cause bacterial mutation. DCAA has a carcinogenic effect on rodents, and an higher hereditary than TCAA. With respect to adverse effects, DCAN leads to mutagenicity in bacterial assays (Oliver 1983), and some of these DBPs (e.g., HANs, HNMs, and HAcAms) have significantly higher cytotoxicity and genotoxicity than the regulated THMs and HAAs (Richardson et al. 2007). Moreover, 1,1-DCP and 1,1,1-TCP have carcinogenic and mutational effects on rats (Bull & Robinson 1986).

Consistent efforts have been made to determine the identities and toxicities of various DBP species and their groups, especially those of THMs, HAAs, and HANs to model their formation and to control their occurrence. Currently, no studies have investigated the monochloramination of co-existing Microcystis aeruginosa and Cyclops metabolites. The objective of this research was to evaluate the formation of selected DBPs during the monochloramination of co-existing Microcystis aeruginosa and Cyclops metabolites, with the dissolution evaluated under various conditions, including the monochloramine dosage, reaction time, pH, temperature, Cl/N, and Microcystis aeruginosa density. The concentration of the co-existing Microcystis aeruginosa and Cyclops metabolites in solution was measured as the total organic carbon (TOC).

Deionized water was used to prepare all solutions. Methanol, acetone and methyl tert-butyl ether (MTBE) were all HPLC grade. The monochloramine solution (500 mg/L) was freshly prepared by mixing a free chlorine solution with an ammonium chloride (NH₄Cl) solution at an initial Cl/N mass ratio of 4/1. Buffer solutions at pH 5, 6, 7, 8, 9, and 10 were prepared with phosphate salts (Tianjin Chemical Plant, China). Calibration standards, internal standards, and surrogate standards for the analyses of THM, nine HAAs (HAA₉), HAN, CH, and TCNM were obtained from Supelco.

Microcystis aeruginosa (blue-green algae, Collection No. HB909) was cultured in 250 mL flasks containing 150 mL of BG11 media under a fluorescent lamp with an automated light/dark cycle of 12 h/12 h in an incubator at 23 °C (LRH-250A, TaiHong, China). Microcystis aeruginosa was cultured for 20 days under these conditions.

Cyclops was initially collected from the Mopanshan Reservoir, which is the largest tributary of the Harbin and main drinking water resource. Cyclops was cultured in aerated 10 L glass aquaria filled with raw water from the reservoir. The aquaria were maintained at a constant temperature (23 °C) and exposed to a consistent photoperiod (12 h light/12 h dark). Cyclops was cultured for 2 days under these conditions.

During the period of the outbreak of Microcystis aeruginosa and Cyclops in summer, the monitoring results showed that the number of Microcystis aeruginosa could reach 10⁹ count/L, while Cyclops reached 200 count/L. For this study, 10⁹ count Microcystis aeruginosa and two hundred active Cyclops were cultured in aerated 25 L glass aquaria filled with deionized water. The aquaria were maintained at a constant temperature (23 °C) and exposed to a consistent photoperiod (14 h light/10 h dark). The samples were cultured for 3 days under these conditions, and then they were centrifuged to obtain the upper liquid. One liter of deionized water was added, and the sample suspensions were filtered through a 0.45 μm membrane, the filtrate was analyzed within a week. The TOC concentration was measured based on standards that were prepared by diluting reagents to 5 mg/L. The dissolved organic carbon (DOC) concentration was unchanged.

The TOC was analyzed on a TOC analyzer (TOC-VCPH, Shimadzu). The monochloramine concentration was measured by DPD/FAS titration (Standard Methods 4500-Cl 1998). Analyses of selected DBPs were carried out on a gas chromatography (GC) (Agilent 7890) with an electron capture detector (ECD), following USEPA methods 551.1 (USEPA 1995) and 552.3 (USEPA 2003). The THM, HAN, haloketone (HK), CH, and TCNM concentrations were measured by a liquid-liquid extraction procedure using MTBE and acid methanol, according to USEPA method 551.1 (USEPA 1995). The column used was an HP-5 fused silica capillary column (30 mm × 0.25 mm I.D. with 0.25 mm film thickness). The GC-ECD operating conditions were as follows: detector temperature 290 °C; injector temperature, 200 °C; injection volume 1 mL; and temperature program, 35 °C for 5 min, ramped to 75 °C at 10 °C/min, held for 5 min, then ramped to 100 °C at 10 °C/min, and then held for 2 min.

For DCAA and TCAA analysis, the samples were pretreated with an extraction/derivatization procedure using MTBE and acidic methanol, according to USEPA Method 552.3 (USEPA 2003). The column used was an HP-5 fused silica capillary column (30 mm × 0.25 mm I.D. with 0.25 mm film thickness). The injector, ECD and GC oven temperature programs for compounds other than HAA9 were as follows: injector temperature, 200 °C; ECD temperature, 290 °C; and oven temperature program, 35 °C for 9 min, ramping to 40 °C at 2 °C/min, holding for 8 min, ramping to 80 °C at 20 °C/min, ramping to 160 °C at 40 °C/min and holding for 4 min.

The GC conditions for HAAs analysis were as follows: injector temperature, 210 °C; ECD temperature, 290 °C; oven temperature program, 30 °C for 20 min, ramping to 40 °C at 1 °C/min, ramping to 205 °C at 20 °C/min and holding for 4 min.

The stock solution containing the co-existing Microcystis aeruginosa and Cyclops metabolites was diluted with deionized water to prepare a testing solution with 5 mg/L as TOC. Monochloramine was added at 10 mg/L to the solution (5 mg/L as TOC), which was buffered at pH 7.0 at a room temperature of 23 ± 1 °C as the baseline condition. The reaction was terminated by Na₂SO₃ prior to DBP extraction. The influencing factors and their tested levels were as follows: monochloramine concentration (2, 4, 8, 10, 20 mg/L as Cl₂), reaction time (6, 12, 24, 48, and 72 h), pH (5, 6, 7, 8, 9, and 10), temperature (10, 20, and 30 °C), Cl/N ratio (1/0, 20/1, 5/1, and 5/4), and Microcystis aeruginosa density (10⁸, 10⁹, and 10¹⁰ count/L).

The formation of DCAA and TCAA increased with increasing pH from 5 to 9, but when the pH was further increased to 10, the concentration of TCAA decreased. The concentration of CH increased with the pH from 5 to 8, but at pH 9 and above, CH was almost undetectable (detection limit of 0.001 μg/L). Monochloramine was the dominant chloramine at pH 7.5 or above, for which the sterilization ability is poor. The hydrolysis of monochloramine to form free chlorine was also affected by pH (Jolley & Carpenter 1983). The pH also affects the stability of unstable DBPs. DCAN, 1,1-DCP, and 1,1,1-TCP can be hydrolysed and decomposed in alkaline conditions (Yang et al. 2007), where the hydrolysis rate is accelerated by the increase in pH (Nikolaou et al. 2000). The hydrolysis rates of these unstable DBPs increase with increasing pH (Xie 2001).

The co-existence of Microcystis aeruginosa and Cyclops metabolites could produce DBP precursors during monochloramine disinfection. Increasing reaction time and monochloramine dosage increased the formation of the relatively stable C-DBPs (e.g., THM, HAA, and CH). The formation of TCNM followed an increasing and then decreasing trend with prolonged reaction time and increased monochloramine dosage. The concentrations of DCAA, and TCAA were highest among all the DBPs (5–8 μg/L), and these compounds deserve further study. The pH affected the formation of DBPs in a different way. TCM was the only species that steadily increased in concentration with increasing pH, the HK concentrations varied in an opposite manner, and other DBPs had maximum concentrations at certain pH values. Higher temperatures enhanced TCM, TCNM, DCAA, and TCAA formation whereas the CH, DCAN, 1,1-DCP, and 1,1,1-TCP concentrations increased between 10 °C and 20 °C, followed by a decrease between 20 °C and 30 °C. The five most common DBPs decreased with a decreasing Cl/N mass ratio, indicating that a lower Cl/N mass ratio can effectively control the amounts of these five DBPs. Finally, the minimum DBP formation was observed from the data with a Microcystis aeruginosa density of 10⁹ count/L.

The project was supported by Foundation item: the National Natural Science Foundation of China (NO.503780262); Supporting Certificate of China Postdoctoral Science Foundation (NO.20070420882); and the National Natural Science Foundation of Heilongjiang Province of China (NO. E200812).