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Algae-induced photodegradation of antibiotics: a review

Lianxue Wei, Haixiao Li, Jinfeng Lu

PII: S0269-7491(20)36277-1
DOI: https://doi.org/10.1016/j.envpol.2020.115589
Reference: ENPO 115589

To appear in: Environmental Pollution

Received Date: 19 May 2020

Revised Date: 21 July 2020
Accepted Date: 1 September 2020

Please cite this article as: Wei, L., Li, H., Lu, J., Algae-induced photodegradation of antibiotics: a review,
Environmental Pollution, https://doi.org/10.1016/j.envpol.2020.115589.

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Graphical Abstract

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1 Algae-induced photodegradation of antibiotics: a review

2 Lianxue Wei1, 2, Haixiao Li1, 2, Jinfeng Lu1, 2, 3

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3 College of Environment Science and Engineering, Nankai University, Tianjin, 300350

4 China
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5 Tianjin Key Laboratory of Environmental Technology for Complex Trans-Media

6 Pollution, Nankai University, Tianjin, 300350 China

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7 Key Laboratory of Pollution Processes and Environmental Criteria (Nankai University),

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8 Ministry of Education, Tianjin 300071, China

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10 Corresponding author:
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11 Jinfeng Lu
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12 E-mail: [email protected] (J. Lu)

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13 Number of tables: 2

14 Number of figures: 4
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15 Abstract

16 Antibiotics are a typical group of pharmaceutical and personal care products (PPCPs)

17 with emerging pollutant effects. The presence of residual antibiotics in the environment is

18 a prominent issue owing to their potential hazards, toxic effects, and persistence. Several

19 treatments have been carried out in aquatic environments in order to eliminate antibiotic

20 residues. Among these, photodegradation is regarded as an environmentally-friendly and

21 efficient option. Indirect photodegradation is the main pathway for the degradation of

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22 residual antibiotics in natural water, as opposed to direct photodegradation. Algae,

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23 working as photosensitizers, play an important role in the indirect photolysis of residual

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antibiotics in natural water bodies. They promote this reaction by secreting extracellular
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25 organic matters (EOMs) and inducing the generation of active species. In order to provide
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26 a thorough understanding of the effects of algae on residual antibiotic degradation in the

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27 environment, this paper comprehensively reviews the latest research regarding algae-

28 induced antibiotic photodegradation. The summary of the different pathways and

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29 photosensitive mechanisms involved in this process show that EOMs are indispensable to
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30 antibiotic photodegradation. The influencing factors of algae-induced photodegradation

31 are also discussed here: these include algae species, antibiotic types, and environmental

32 variables such as light source, ferric ion presence, temperature, and ultrasound treatment.

33 Based on the review of existing literature, this paper also considers several pathways for

34 the future study of algae-induced antibiotic photodegradation.

35 Keywords: antibiotics, photodegradation, algae, extracellular organic matters,

36 photosensitizers

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37 Introduction

38 In recent years, residual antibiotics have drawn much attention due to their harmful

39 influence on the ecological environment and human health (Guo et al., 2016; Li and Hu,

40 2016; Baena-Nogueras et al., 2017). In addition to medical use, antibiotics are also used

41 as pesticides and growth promoters in agriculture and aquaculture, and to treat livestock

42 (Li and Hu, 2016; Tian et al., 2018). Given this continuous input into environment,

43 antibiotics cannot be completely metabolized by plants or animals (Sarmah et al., 2006;

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44 Xu et al., 2011; Wang et al., 2017). At the same time, the residual antibiotics are hard to

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45 be degraded through self-degradation and biosorption in nature (Guo and Chen, 2015;

46
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Fabrice et al., 2019). Consequently, both the continuous input and slow removal of
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47 antibiotics lead to their long-term presence in the environment (Monteiro and Boxall,
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48 2010; Bonvin et al., 2012). Antibiotic residues have been detected in many aquatic
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49 environments (Peng et al., 2011; Wammer et al., 2013; Wang et al., 2017). Yiruhan et al.

50 (2010) reported fluoroquinolones in tap water from Guangzhou and Macao, in

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51 concentrations of 1.0–679.7 ng L-1 and 2.0–37.0 ng L-1, respectively. Higher residual

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52 antibiotic concentrations of 652–5770 ng g-1 in the sediments of Chinese rivers have been

53 reported in the studies of Peng et al. (2008, 2009, and 2011). In addition, Li et al. (2013)

54 found quinolone concentrations of up to 29647 mg kg-1 in domestic sludge.

55 The long-term presence of residual antibiotics in the environment can potentially

56 give rise to hazards such as ecological toxicity and the occurrence of resistant genes. For

57 example, Dong et al. (2012) found that tetracycline exposure induced the breakage of

58 DNA in earthworms, changing enzymatic activity. Similar phenomena occur in plants,

59 for example, the chloroplastic and mitochondrial protein synthesis of two species of ferns

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60 is affected by residual antibiotics such as tetracyclines and fluoroquinolones (Kümmerer,

61 2011). Additionally, numerous bacteria species with antibiotic resistance genes have been

62 reported in soil and aquatic environment due to anthropogenic sources (Erik et al., 2011;

63 Bonvin et al., 2012; Gothwal and Shashidhar, 2015) like fertilizing the soil with

64 antibiotic-contaminated manure from farms and the release of pharmaceutical wastewater

65 (Choi et al., 2020) as well as the presence in aquacultural habitats like the shrimp farm

66 ponds (Zhou et al., 2020). The overuse of antibiotics induces the antibiotic resistance

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67 genes in animals and further leads to horizontal gene transfer to bacterial pathogens

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68 (Yang et al., 2013; Zhu et al., 2013; Luo et al., 2017). Generally, DNA is still contained

69
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in faeces and residues of animals with antibiotic resistance genes, which can easily flow
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70 into the environment. Then, exogenous DNA could be transferred to bacterium through
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71 different mechanisms (bacterial conjugation, natural transformation, transduction) (Boto

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72 et al., 2019; Sun, 2018). It was also convinced by the finding that farm antibiotic use

73 increased the frequency of horizontal gene transfer and was closely related to the growth
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74 and spread of antibiotic resistance genes in bacterial pathogens (Levy, 1978; Gillings and
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75 stokes, 2012). The existence of such resistance genes alters microbial communities (Chen

76 et al., 2016; Li and Hu, 2016). Moreover, the antibiotic resistance of bacterial pathogens

77 may prevent the effective treatment of some diseases and cause damage to human health

78 (Magdalena et al., 2019). In this context, finding efficient and environmentally-friendly

79 solutions to remove residual antibiotics from the environment is necessary. In general,

80 sorption, indirect photodegradation (or indirect photolysis), oxidation, and biodegradation

81 are the most common methods of eliminating residual antibiotics in the environment

82 (Poiger et al., 1999; Knapp et al., 2005; Zwiener, 2007; Li and Zhang, 2010).

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 83 Sorption treatment of residual antibiotics has been studied by Westerhoff et al.

84 (2005), they found that the sorption of powdered activated carbon could contribute to the

85 elimination of residual antibiotics in river water: a removal rate of 49–99% was identified

86 for residual antibiotic concentrations varying from 10 mg L-1 to 20 mg L-1 after a contact

87 time of 4 h. However, sorption can only transfer pollutants, it cannot completely

88 eliminate them (Homem and Lucia, 2011). Thus, the post-treatment is required to

89 eliminate the adsorbed antibiotics from the adsorbent materials and the recovery of

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90 adsorbent is also a problem yet to be solved (Adams et al., 2002).

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91 Biodegradation of residual antibiotics requires harsh conditions as most antibiotics

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are difficult to biodegrade due to their pharmacological stability (Fabrice et al., 2019) or
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93 resistance (Khetan and Collins, 2007; Xu et al., 2011). This method is used in
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94 combination with other processes to remove residual antibiotics, especially in the

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95 activated sludge processes of wastewater treatment plants (Li and Zhang, 2010).

96 However, biodegradation is not suitable for wastewater with high concentrations of

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97 residual antibiotics as this would inhibit the activities of microorganisms (Britto and
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98 Rangel, 2008; Homem and Lucia, 2011). Furthermore, the biodegradation method can

99 also lead to resistant genes.

100 Oxidation was successfully used for the removal of sulfonamides and trimethoprim

101 by Gothwal and Shashidhar (2015): ozone treatment at a dosage of 7.1 mg L-1 led to a

102 removal rate of over 95%. Even though oxidation techniques such as ozonation are

103 efficient, they produce some by-products which may be ecologically toxic (Le-Minh et al.,

104 2010; Gothwal and Shashidhar, 2015). Furthermore, oxidation treatment requires

105 expensive equipment as well as large amounts of energy (Homem and Lucia, 2011). In

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106 addition, ozonation may make residual antibiotics more resistant to further ozonation (Le-

107 Minh et al., 2010).

108 In contrast, photodegradation is one of the main transformation processes of residual

109 antibiotics in natural water environments, especially in surface layers (Doll et al., 2003;

110 Dabic et al., 2019). Most antibiotics are highly sensitive to light (Bonvin et al., 2013;

111 Baena-Nogueras et al., 2017; Tian et al., 2018). Antibiotics can be directly activated to

112 degrade or break up when absorbing light, this is direct photodegradation (Zepp and

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113 Cline, 1977; Wammer et al., 2013; Tian et al., 2018). While indirect photodegradation

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114 depends on oxidant species (e.g., hydroxyl radicals (·OH), singlet oxygen (1O2),

115
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superoxide (O2-· ), hydrogen peroxide (H2O2), peroxyl radicals (·OOR)) and triplet
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116 excited-state dissolved organic matter (3DOM*) which produced by photosensitizers
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117 under light (Li and Hu, 2016; Tian et al., 2018). Involving active species could greatly
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118 shorten the half-lives of residual antibiotics and increase their degradation rates. The

119 reaction rate of indirect photolysis is much faster than direct photolysis and plays a
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120 dominant role in the removal of organic chemicals (Xu et al., 2011; Tian et al., 2018). In
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121 natural water bodies, algae are effective photosensitizers that can generate reactive

122 oxygen species (ROS) under irradiation (Zepp and Schlotzhauer, 1983). Thus, studies

123 have focused on the contribution of algae to the efficiency and effectiveness of residual

124 antibiotic removal in aquatic environments. Tian et al. (2018) found that the presence of

125 Chlorella vulgaris under irradiation led to a chlortetracycline (CTC) removal rate of close

126 to 96%. Since no other chemicals were added to this process, photodegradation induced

127 by algae would therefore be the reason for the high removal rate. Additionally, Tian et al.

128 (2019) reported that the algae C. vulgaris, Microcystis aeruginosa, and Scenedesmus

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129 promote the photodegradation rate of CTC, while Zhang et al. (2012) noted that

130 norfloxacin degradation rate in a solution of C. vulgaris under light is three times faster

131 than that without algae. The algae induced photodegradation has also been adapted in

132 wastewater treatment. For instance, Guo and Chen. (2015) has applied the approach of

133 the alga-activated sludge combined system to achieve the excellent performance both in

134 residual antibiotic (cefradine, cephalexin, ceftazidime and cefixime) removal and in

135 activation of sludge. The antibiotic degradation was closely related to the generation of

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136 ROS such as ·OH. The residual antibiotics could be degraded into small molecular

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137 substance by ROS. Moreover, Villar-Navarro et al, (2018) have performed an experiment

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at pilot scale to explore the efficiency of microalgae biotechnology system on
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139 pharmaceutical removal in a WWTP. Results showed that the efficiency of microalgae
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140 treatment on drugs (>70%) was higher than that of the activated sludge process. Thus,
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141 algae-based treatment is a promising application for real wastewater treatment.

142 In recent years, research into the indirect photolysis of residual antibiotics induced
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143 by algae has escalated, with the number of published manuscripts on the topic steadily
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144 increasing from 2009 to 2019 (Fig. 1). However, the results of photodegradation on

145 different antibiotics and using different algae species are inconsistent, with the unclear

146 effects of environmental conditions further confusing the matter. The mechanisms behind

147 photodegradation efficiency should be discussed in a comprehensive way. Thus, a review

148 of the current literature would contribute to a better understanding of algae-induced

149 photodegradation of antibiotics and provide potential pathways for future study. In this

150 review, the degradation pathways and mechanisms of algae-induced photodegradation are

151 introduced. Then, the effects of algae species, antibiotic species, and other environmental

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152 conditions on photodegradation efficiency are discussed to give a comprehensive

153 understanding of the topic.

154 1. Mechanisms of indirect antibiotic photodegradation induced by algae

155 Algae-induced photodegradation of residual antibiotics is a type of indirect

156 photodegradation that depends on the active oxidizing agents generated by

157 photosensitizers. As previously reported, many substances contained in algae solutions

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158 can act as photosensitizers such as chlorophyll, enzymes, and extracellular organic

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159 matters (EOMs; Zhang et al., 2012; Tian et al., 2018).

160 In general, algal organic matters (AOMs) can be divided into EOMs and

161
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intracellular organic matters (IOMs; Li et al., 2012; Doorae et al., 2018). EOMs are
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162 formed by extracellular metabolic excretion of algae, whereas IOMs arise from
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163 intracellular autolysis of algal cells. IOMs account for 80% of the total AOM (Henderson
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164 et al., 2008; Doorae et al., 2018). However, EOMs tend to play the dominant role during

165 photodegradation by producing active species under irradiation (Tenorio et al., 2017;
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166 Tian et al., 2018; Tian et al., 2019). This might be because EOMs are composed of
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167 proteins, carbohydrates, and humic-like substance (Henderson et al., 2008; Li et al., 2012;

168 Qu et al., 2012). Humic substances can act as photosensitizer and greatly affect the

169 photodegradation of residual antibiotics (Ma et al., 2010; Li and Hu, 2016). It was

170 confirmed that enzymatic proteins account for half of the total proteins in algal EOMs

171 (Rao et al., 2020), which played minor role in antibiotic degradation (Tian et al., 2018).

172 However, the proteins involved in photosynthesis might be seen as photosensitizer. And

173 there is a lack of research on whether carbohydrates could be used as photosensitizer at

174 present. Furthermore, the dissolved parts of EOMs contribute significantly to

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175 photodegradation. For example, humic and fulvic dissolved organic matters (DOMs) is

176 photochemically reactive and beneficial to the photolysis process (Zuo et al., 1996;

177 Franke and Franke, 1999). Zhang et al. (2012) found that the DOMs released by algae

178 under irradiation could act as a photosensitizer in the photodegradation of norfloxacin.

179 Furthermore, Tranvik and Kocalj (1998) noted that DOMs had a strong capacity to

180 absorb photons; it is therefore reasonable to assume that DOMs from algal EOMs are the

181 main driver for induced antibiotic photodegradation.

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182 Algal EOMs initiate antibiotic photodegradation through generating reactive

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183 oxidizing species (ROS). When comparing other ROS such ⋅ OH and 1O2, the quenching

184
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experiments of Tian et al. (2019) showed that triplet excited-state extracellular organic
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185 matter (3EOM*) accounted for 90% of the photolysis process induced by EOMs. The rate
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186 constant for the photo-reaction of 3EOM* with chlortetracycline was 3.17×109 M-1S-1,
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187 which was much faster than that of 1O2 (1.5×106 M-1S-1) (Tian et al., 2018). It implies the
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188 active degree of active species. However, the rate constant is closely related to the steady
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189 state concentration of ROS according to Mostofa et al. (2013). Thus, playing a minor role

190 in the photo-reaction does not mean that this kind of ROS has low activity. It is also

191 possible that its steady state concentration is really low. Moreover, according to the

192 experiments by Ma et al. (2010), which used NaBH4 to selectively reduce carbonyl-

193 containing compounds, EOMs with more carbonyl compounds may have higher photo-

194 sensitivity. This is supported by Tian et al. (2019). EOMs containing more carbonyl

195 compounds are likely to generate more 3EOM*. This indicates that 3EOM* plays the

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196 dominant role in photo-reactivity of algae, although the performance of 3EOM* from

197 different algae species may vary.

198 In promoting the indirect photodegradation of residual antibiotics, IOMs play only a

199 minor role. One reason for this may be that the ROS generated by IOMs cannot pass

200 through the cell membrane to make contact with antibiotics. In addition, Pivokonsky et al.

201 (2014) reported that algal IOMs contained more peptides and proteins than EOMs, these

202 are less photochemically reactive than humic substances. However, chlorophyll found in

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203 IOMs may work as photosensitizers. Algae cells are rich in chlorophyll, this substance is

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204 also sensitive to light and can induce the generation of reactive ROS under irradiation

205
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(Liu et al., 2003; Edreva, 2005). One of the main components of chlorophyll, the
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206 substituted porphyrin ring, plays an important role in absorbing energy from light sources
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207 (Rebelo et al., 2007; Luo et al., 2015). Therefore, chlorophyll is activated after the
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208 absorption of light energy (Ge and Deng, 2015) and either directly reacts with pollutants

209 or generates active species. Peng et al. (2006) reported that chloroplasts from C. vulgaris
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210 could form oxygen species when subjected to light, thus promoting bisphenol A (BPA)
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211 degradation. Moreover, Zhang et al. (2012) found that chlorophyll was an important

212 substance in the photoreactions of norfloxacin. Tian et al. (2018) also found that

213 chlorophyll accelerated the photodegradation of chlortetracycline, although the removal

214 rate was only enhanced by 3%.

215 As catalysts, enzymes in both algal EOMs and IOMs may be another driver for

216 induced antibiotic photodegradation. However, their effects on antibiotic

217 photodegradation are not currently evident. Tian et al. (2018) found that the effects of

218 enzymes and algal chlorophyll on the degradation of chlortetracycline were insignificant

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220 presence of algae are summarized in Fig. 2.

221 2. Effects of algae species on photodegradation

222 The efficiency of algae-induced antibiotic photolysis may be affected by which

223 algae species is used. Ge and Deng. (2015) explored the efficiency of two algae species

224 (Platymonas subcordiformis and Isochrysis galbana) in the photodegradation of two

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225 fluoroquinolone antibiotics (enrofloxacin and ciprofloxacin hydrochloride). Both algae

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226 had promoting effects on the photodegradation of those two antibiotics. However, P.

227 subcordiformis improved the average photolysis rate to 76.1%, whereas the rate under I.

228
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galbana treatment was 68.7%. Moreover, Tian et al. (2019) studied the indirect
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229 photolysis efficiency of three different algae species on CTC, namely M. aeruginosa
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230 (MA), Scenedesmus meyen (SM) and C. vulgaris (CV). The experiments were carried out
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231 under the same environmental conditions with a xenon lamp as a light source.

232 Photosensitizers involved in the reaction (EOMs) were extracted from these three algae
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233 species. Results showed that all extracted EOMs promoted the degradation of CTC to
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234 varying degrees (CV-EOMs > SM-EOMs > MA-EOMs). Another study showed that the

235 cefradine (CED) removal rate of Chlamydomonas reinhardtii was 100% under ultraviolet

236 light while that of Chlorella pyrenoidosa was 77.99% (Du et al., 2015; Jiang et al., 2019).

237 Thus, the performance of algae-induced antibiotic photodegradation is species specific.

238 This specificity is likely due to two reasons. Firstly, the ROS generated by different

239 algae species varies. Tian et al. (2019) found that the EOMs generated by C. vulgaris and

240 S. meyen had a higher yield of 3EOMs* than that of M. aeruginosa since their EOMs

241 contained more carbonyl compounds. As 3EOMs* are the major contributors to the

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242 photodegradation process induced by algal EOMs, C. vulgaris and S. meyen will favour

243 antibiotic photodegradation more than M. aeruginosa. Therefore, if an algae species

244 excretes EOMs containing more carbonyl compounds, such algae would be more

245 favourable for the photodegradation of residual antibiotics.

246 Secondly, the effects of algae on the photodegradation rate of residual antibiotics are

247 also largely related to algae concentration, which may affect photoproduct yield. As

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248 reported by Zhang et al. (2012), the higher concentration of C. vulgaris (1.7×106-1.7×

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249 107 cell L-1) led to a faster degradation rate of norfloxacin. This could be explained by the

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fact that more algae produce more organic matter to serve as photosensitizers for
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251 photodegradation. The algae cell density is known to be highly related to algae growth

252 stages, hence choosing the algae of proper growth stage is important for improving
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253 antibiotic photodegradation. In most cases, the algae are used in experiments when the
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254 cells grow in logarithmic phase (Wang et al., 2007; Tian et al., 2018; Jiang et al., 2019).
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255 However, residual antibiotics can also affect the growth of algae, even causing toxic
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256 effects in some cases (Chen and Guo, 2012; Liu et al., 2012; Shang et al., 2015). For

257 example, Schlegel et al. (1998) used antibiotics against green algae while Du et al. (2018)

258 found that three antibiotics (cefradine, norfloxacin, and amoxicillin) had inhibiting effects

259 on M. aeruginosa growth. This is likely due to cefradine and amoxicillin limiting algal

260 cell wall synthesis, and norfloxacin directly reducing algal cell size. Liu et al. (2014) also

261 reported that a mixture of spiramycin and amoxicillin was able to inhibit the growth of M.

262 aeruginosa, depending on the ratio and the concentration. Additionally, Carusso et al.

263 (2018) found that chlortetracycline was more toxic than enrofloxacin to

264 Pseudokirchneriella subcapitata and Ankistrodesmus fusiformis. Hence, when using

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265 algae to stimulate antibiotic photodegradation, it is important to ensure that algae growth

266 is not inhibited by the antibiotics themselves. It is essential to choose the right algae

267 species for antibiotic photodegradation to avoid such inhibiting effects.

268 The decrease in the antibiotic degradation rate as algal concentration increases, up to

269 a certain level, may be due to the screening of light by algae. Jiang et al. (2019) explored

270 the effects of Chlamydomonas reinhardtii density on cefradine degradation and found

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271 that the removal rate of cefradine increased with increasing algal density but declined

with cell densities above 50 × 104 cells mL-1, likely due to shading effects. Thus, an

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appropriate algae concentration is required for optimal degradation efficiency.
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274 3. Effects of antibiotic type on photodegradation
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275 Algae and their secretions can absorb the light radiation of a specific wavelength

276 and then generate ROS through a series of reactions (Qu et al., 2012; Tenorio et al., 2017;
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277 Sandvik et al., 2000), which favour the photodegradation of residual antibiotics. However,
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278 different antibiotics respond differently to algal photodegradation. In Table 1, the

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279 removal rates of several antibiotics under algae induced photodegradation are listed.

280 Regardless of other conditions, the CTC tends to be the most easily degraded by algal

281 photodegradation with an average removal rate of 88%. This is followed by Enrofloxacin

282 (ENR; 79.9%), Ciprofloxacin hydrochloride (72.3%), Norfloxacin (NOF; 47.0%), and

283 Cefradine (CRD; 42.66%).

284 Chlortetracycline belongs to the group of tetracycline antibiotics, which are ranked

285 second in production and usage all around the world (Levy, 2002). The main function of

286 CTC is to promote the growth of plants or animals. As early as 2009, it was reported that

287 their residues in the environment have led to the generation of tetracycline resistance

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288 genes (Zhang et al., 2009). Currently, the widespread occurrence of CTC in the effluent

289 of wastewater treatment plants (WWTPs) has been of concern (Shetty, 2002).

290 Chlortetracycline is easier to degrade than other antibiotics that remain in the

291 environment because it contains more functional groups that are prone to photo-oxidative

292 reactions, such as -OH and -NH2 (Tian et al., 2018). The increase of the degradation rate

293 of CTC in the presence of C. vulgaris (Tian et al., 2018), is mainly due to the 3EOM*

294 generated by algae under irradiation, although the small amount of ROS, such as

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295 O2, ·OH, produced in the process also contributed to some extent (Tian et al., 2018; Tian

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296 et al., 2019). According to the mechanisms mentioned above, residual antibiotics

297
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generally have toxic effects on algae cells, further inhibiting the generation of ROS. In
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298 contrast, the extraction of EOMs from algae is not affected by residual antibiotics. This is
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299 one of the reasons behind the higher removal rate of CTC. The possible degraded
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300 mechanisms of CTC are illustrated in Fig. 3(a).

301 Norfloxacin, with the chemical formula 1-ethyl-6-fluoro-1,4-dihydro-4-oxo-7-

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302 (piperazinyl) quinoline-3-carboxylic acid, is a typical fluoroquinolone in that it is a

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303 broad-spectrum antibacterial agent (Al-Tamrah et al., 2015). It is widely used in human

304 and veterinary medicine, especially in the treatment of urinary tract infections (Córdoba-

305 Diaz et al., 1998; Feng et al., 2017). This antibiotic is consumed in large quantities every

306 year; according to the reports, over 500 tons were consumed in 2003 (Huang et al., 2017;

307 Jing et al., 2019). This extensive use of NOF makes it the second most detected

308 fluoroquinolone in aquatic environments (Zhang et al., 2012; Rahman et al., 2004). To

309 explore the NOF degradation efficiency induced by algae, Zhang et al. (2012) conducted

310 a series of experiments using C. vulgaris. As shown in Table 1, the reaction system

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311 containing algae alone greatly improved the photolysis rate, which was approximately

312 three times faster than the control group. But this removal rate was still much lower than

313 that of CTC. This may be because, unlike CTC, the structure of NOF lacks the functional

314 groups that are prone to photo-oxidation reactions. This is supported by the observation

315 that the direct photodegradation of CTC reached a rate of 48% under irradiation (Tian et

316 al., 2018), while NOF was stable and its direct photodegradation had little effect (Lv et

317 al., 2017). During the photodegradation of NOF, ·OH is the dominant active species,

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318 being involved in the reaction of hydroxylation, decarboxylation, and piperazinyl ring

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319 cleavage during the degradation process (Lv et al., 2017; Zhou et al., 2017). However, in

320
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algae solution under irradiation, the generation of ·OH is mainly induced by the
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321 absorption of light by photosensitizers such as humic and fulvic DOMs or chlorophyll
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322 (Zhang et al., 2012; Edreva., 2005; Liu et al., 2003), and the yield of ·OH was relatively
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323 low in the process of algae-induced photodegradation according to Tian et al., (2018). In

324 addition, NOF can cause irreversible damage to algae cells (Du et al., 2018), potentially
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325 decreasing the efficiency of NOF degradation by C. vulgaris. The possible degradation
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326 mechanisms for NOF are illustrated in Fig. 3(b).

327 Variation in antibiotic structure, which makes them recalcitrant and persistent in

328 natural water bodies, is the main reason behind their different degradation rates.

329 According to Tian et al., (2018), -OH and -NH2 are sensitive to light. Antibiotics

330 containing these functional groups can therefore be easily degraded. Furthermore,

331 antibiotics can be more easily degraded under irradiation if they contain aromatic rings

332 and heteroatoms (Hao et al., 2019; Kim et al., 2009). Table 2 details the structure of

333 different antibiotics mentioned in the cited literature.

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334 4. Other treatments

335 In addition to algae species and antibiotic type, algae-induced antibiotic

336 photodegradation is also influenced by other treatments such as light source selection, pH

337 adjustment, temperature conditioning, ultrasound treatment, and the application of ferric

338 ions (Fe3+).

339 The energy from different light sources undoubtedly influences algae-induced

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340 photodegradation. Jiang et al. (2019) found that Chlamydomonas reinhardtii-induced

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341 photodegradation of CED had a removal rate of 14.72 ± 0.93%. However, CED was

342

343
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completely degraded under ultraviolet light with the same conditions. The efficiency of

combining algae and ultraviolet light is likely due to high-energy light sources favouring
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344 the generation of ROS such as 3EOM*.
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345 The environment pH would influence algae-induced antibiotic photodegradation by

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346 firstly affecting algae secretions. According to Zhang et al. (2012), NOF is easier to
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347 degrade in weak acidic environments, because more dissolved organic matters are
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348 released by algae under these conditions. On the other hand, pH may affect the properties

349 of antibiotics like the protonation state, which has an impact on antibiotic optical

350 properties and reactivity (Boreen et al., 2004). For instance, as amphoteric substrates, the

351 species of NOF in solution would alter depending on pH (Takács-Novák et al., 1990;

352 Elisa et al., 2010). It would probably be another reason why NOF could be more

353 photodegraded in weak pH environment. The effect of pH on antibiotic photodegradation

354 has also been proved by the study of Linke et al (2010) that the photolysis rate constants

355 of sarafloxacin (SAR) and gatifloxacin (GAT) increased first and then decreased as pH

356 increasing with a peak value at pH=8. According to the pKa1 and pKa2 showed in Table 2,

16
357 the form of zwitter ions were dominant at pH=8 for SAR and GAT, which photodegraded

358 faster than other forms. In short, pH could influence algae-induced antibiotic

359 photodegradation mainly by affecting algae secretions and antibiotic forms. The octano-

360 water partition coefficients (logKow) of typical antibiotics have been presented in Table

361 2, and it shows the degree of mixing between algae and antibiotics.

362 Temperature is another influencing factor for the algae induced photodegradation of

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363 residual antibiotics. The increase of temperature (<45 ) is beneficial for the photolysis of

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364 antibiotics (Zhang et al., 2012). It can affect the algae growth and the amount of algae

365

366
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secretion. A suitable temperature is beneficial to algae growth. According to the previous

reports, improving the temperature in the optimal range will greatly increase the
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367 metabolic rate of algae and promote algae growth (Hong et al., 2019; Zachleder et al.,
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368 2019). Different algae have different optimal temperature for growing, for example,
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369 Chlamydomonas are prefer to grow at 35 . However, the growth temperature is

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370 generally not higher than 42 , which may lead to the cell cycle arrest according to

371 Zachleder et al, (2019). Moreover, as the algae grow faster with the increasing
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372 temperature, more secretion would be released accordingly (Zhang et al., 2012). It would

373 be beneficial to promote the photo-reaction process through generating amount of active

374 species.

375 The performance of algae induced photodegradation is also affected by the

376 pretreatment process like ultrasound. The ultrasound pretreatment can promote the

377 reaction by making a more uniform distribution of antibiotics and algae (Huang et al.,

378 2019; Zhang et al., 2009). Moreover, Zhang et al., (2012) found that ultrasound treatment

379 promotes the degradation of NOF in algae solution. It was because that ultrasound broke

17
380 apart the algae cells, releasing low molecular weight organic in the reaction system. If

381 oxygen is bubbling into the solution, 1O2 would be produced due to the transformation of

382 O2 with ultrasound energy, which would also favor the antibiotic degradation (Li et al.,

383 2020). However, this process would be blocked due to the degassing of dissolved oxygen

384 by ultrasound (Laugier et al., 2008).

385 Ferric ion application is especially important in algae-induced antibiotic

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386 photodegradation. One of the most common metal ions in natural water environments is

Fe3+. This ion is able to form complexes with carboxylic acid, making Fe( )-carboxylate

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387

388

389
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complexes. Additionally, it is more photoactive in generating free radicals under

irradiation (Goldberg et al., 1993; Wang et al., 2009; Li et al., 2010; Zhang et al., 2012).
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390 Previous research also found that the complexes formed by Fe3+ and pyruvic acid were
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391 the sources of ·OH (Wang et al., 2006). Wang et al. (2009) explored the mechanisms of
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392 pollutant degradation influenced by Fe( )-carboxylates especially with oxygen

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393 introduced into the solution. During photo-reaction, the Fe( )-carboxylate complexes

394 could be photolyzed easily and the introduced oxygen promote this process. Fe3+ was
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395 firstly reduced to Fe2+, after a series of internal cross reactions, H2O2 was produced.

396 Finally, ·OH was generated due to the Fenton reaction. This process has been confirmed

397 by several reports (Zuo and Hoigne, 1993; Wu et al., 2004). Interestingly enough, a

398 competition between Fe2+ and organic pollutants for ·OH was existing in the system, and

399 results showed that the reaction between ·OH and organic pollutants was dominant. Thus,

400 the combination of algae and Fe3+ is a feasible and alternative method of promoting the

401 degradation of antibiotics. The compounds containing carboxyl structures released by

402 algae may react with ferric ion to improve the antibiotic photolysis rate. This statement is

18
403 supported by the observation that 80.3% of NOF was removed in an algae solution

404 containing Fe3+ after 60 min irradiation, which was much faster than that without Fe3+

405 (Zhang et al., 2012). Furthermore, another study showed that a higher enrofloxacin and

406 ciprofloxacin hydrochloride removal efficiency is observed in Fe (III)-algae suspensions

407 (Ge and Deng, 2015).

408 5. Conclusion

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409 This review provides a comprehensive overview of the indirect photodegradation of

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410 antibiotics induced by algae. The efficiency of this process is determined by the chemical

411 structures and properties of antibiotics and algae, as well as other environmental

412
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conditions, although chemical characteristics play the dominant role in determining
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413 efficiency. The chemical structure of antibiotics influences their persistence. Antibiotics
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414 containing photosensitive functional groups such as -OH and -NH2 are easier to degrade
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415 under irradiation. The chemical structure of EOMs extracted from algae influence their

416 photosensitive activity. The dominant substances in the photodegradation reaction have
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417 been found to be EOMs; EOMs that contain more carbonyl compounds release more
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3
418 EOMs*, which improves antibiotic removal. However, residual antibiotics generally

419 have toxic effects on algae cells, inhibiting algal secretion yield. As such, extracting

420 EOMs from algae is considered a good method to avoid the adverse effects of residual

421 antibiotics on algae cells. In algae induced photodegradation, EOMs-generated ROS such

422 as ·OH, 1O2, and 3EOM* accelerate the reaction rates of residual antibiotics. Among
3
423 these ROS, EOM* makes the greatest contribution (90%) to photodegradation.

424 Moreover, the addition of Fe( Ⅲ ) and the use of ultraviolet light further improve

425 antibiotic degradation rates.

19
426 Algae that are found extensively in aquatic environments show great potential for

427 removing pharmaceutical contaminants. To facilitate the application of algae for

428 environmental purposes, several issues need to be investigated further. Various

429 influencing factors lead to complex interactions between algae and antibiotics. Thus, the

430 appropriate selection of algae species and antibiotic type is worth studying. After the

431 treatment of residual antibiotics by algae, antibiotic resistance genes may transfer and

432 accumulate. However, EOMs extracted from algae can avoid such negative effects; it is

of
433 therefore important to identify the specific substance which is dominant in EOMs. Such a

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434 substance could be used as the adjuvant for the photodegradation of residual antibiotics.

435
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The scientific literature suggests that the contribution of IOMs to antibiotic degradation is
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436 minor, although IOMs account for most AOMs. When algae cells are broken, IOMs
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437 would be released; release would also occur through the autolysis of dead algae cells. The
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438 effect of released IOMs on pollutants under irradiation should therefore be explored

439 further. Algae also play an important role in the treatment of antibiotic residues in
ur

440 wastewater. However, the appropriate method for the addition of algae to the treatment
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441 process should be considered; it is possible that addition of a solution of EOMs extracted

442 from algae would be an appropriate method in this context. It is notable that some

443 antibiotics are difficult to degrade through photodegradation catalysed by algae. Thus, the

444 combination of algae and other pre-treatments such as pre-oxidation should be considered

445 in order to improve degradation efficiency. It is also important to note that some by-

446 products may be produced after algae treatment on residual antibiotics. Studying the

447 composition and the toxicity of such by-products is therefore vitally important.

448 Furthermore, the actual aquatic environment is much more complex than the laboratory

20
449 environment, leading to uncertainty regarding the efficiency of algae in degrading

450 pollutants. The complex substances existing in natural water bodies that have synergistic

451 or antagonistic relationships with algae should therefore also be considered in future

452 research.

453 Conflicts of interest

454 The authors declare no conflicts of interest.

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455 Acknowledgements

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456 This work was financially supported by National Natural Science Foundation of

457 -p
China (Grant No. 51878357); and Major Science and Technology Program for Water
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458 Pollution Control and Treatment of China (2018ZX07110-007).
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767 006-0818-2.

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 20 100

the keywords "photodegradation+antibiotics"

18 90

Numbers of publications containing

Numbers of publications containing
the keywords "algae +antibiotics"

16 80

14 70

12 60

10 50

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8 40

6 30

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4 20

2
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0 0
09

10

11

12

13

14

15

16

17

18

19

20
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20

20

20

20

20

20

20

20

20

20

20

20
768
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769 Fig. 1. Evolution of the number of publications in indexed journals containing the
770 keywords “algae + antibiotics” and “photodegradation + antibiotics” between 2009 and
771 2019.
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772
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773

774
775
776 Fig. 2. The possible schematic for mechanism of the degradation of antibiotics in alga-
777 containing water with irradiation.

33
778
779

780 Fig. 3(a). The possible schematic for mechanism of the degradation of CTC in alga-
containing water with irradiation.

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781

782

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786
787
788 Fig. 3(b). The possible schematic for mechanism of the degradation of NOF in alga-
789 containing water with irradiation.

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 Table 1 The parameters and removal rates of representative drugs

Light Surface light

No. Reference Drugs Algae Concentrations pH Others Removal rates
sources intensity

Chlorella vulgaris; 95%

f
Extracellular

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Tian et Chlortetracycline Scenedesmus meyen; 87%
1 organic matters xenon lamp 30.2mW/cm2 6.8 ——

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al.(2019) (10μM) Microcystis
(4mgC/L) 73.8%

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aeruginosa

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Tian et Chlortetracycline Algae concentration
2 Chlorella vulgaris; xenon lamp 0.89mW/cm2 6.8 —— 96%

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9 -1
al.(2018) (10μM) (2.0×10 cells L )

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ultraviolet 100%

Jiang et Cefradine
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Chlamydomonas Algae concentration
sunlight 32±2.5%
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3 simulated —— 9.0 ——
al.(2019) (5mg/L) renhardtii (1.0×106cells mL-1) 14.72±0.93%
natural light
fluorescent
13.69±1.07%
light
Chlorella Algae concentration —— 17.56%
4 Du et al.(2015) Cefradine 6 -1
—— — ——
pyrenoidosa (10.0×10 cells mL ) ultraviolet 77.99%
5 Ge and Platymonas Algae concentration high- Fe( ) 92%
—— —
algae 81%

35
 Deng.(2015) Enrofloxacin; subcordiformis 3.5×108cells L-1 pressure Fe( )+algae 83%
mercury Fe( ) 74%
Isochrysis galbana 2.08×106cells L-1 algae 71%
lamp
Fe( )+algae 78.5%
Platymonas Fe( ) 80%
3.5×108cells L-1 algae 71.2%
Ciprofloxacin subcordiformis Fe( )+algae 84.2%

f
hydrochloride Fe( ) 55%

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Isochrysis galbana 2.08×106cells L-1 algae 66.4%

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Fe( )+algae 77%
Fe( ) 24.7%

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Zhang et Norfloxacin Algae concentration
6 Chlorella vulgaris Ultraviolet —— 5.0 algae 35.9%

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al.(2012) (20mg/L) (1.7×108cells mL-1)
Fe( )+algae 80.3%

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 Table 2 Physico-chemical properties of representative antibiotics

Therapeutic Molecular Molecular

NO. Reference Drugs CAS Reg.No. Chemical Structure pKa logKow
Class Formula Mass

o f
Tian et al.

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(2018);
1 chlortetracycline tetracyclines 57-62-5 C22H23ClN2O8 478.880 3.3 -0.62

-p
Mariusz et
al. (2019)

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na
Jiang et al. 2.63
2 cerfradine cephalosporins 38821-53-3 C16H19N3O4S 349.405 0.41
(2019) 7.27
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Wammer et
al.(2013); 6.11
3 norfloxacin fluoroquinolones 70458-96-7 C16H18FN3O3 319.331 -1.03
Mariusz et 9.11
al. (2019)

37
 6.2
enrofloxacin 93106-60-6 C19H22FN3O3 359.400 0.7
7.9

f
6.2

o
ciprofloxacin 85721-33-1 C17H18FN3O3 331.342 0.28
8.8

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-p
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Linke et al.
5.9
(2010); sarafloxacin 98105-99-8 C20H17F2N3O3 385.36 ——
4 fluoroquinolones 8.9

na
Mariusz et
al. (2019)
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6.0
gatifloxacin 112811-59-3 C19H22FN3O4 375.394 ——
9.2

38
 1

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Highlights

• The degradation of antibiotics in natural water depends on indirect photodegradation

induced by algae.

• 3
EOM* plays the dominant role in the indirect photodegradation of antibiotics.

• The degradation efficiency of antibiotics is related to algae species, antibiotic types and

their interactions.

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• The combination of algae and Fe3+ is a feasible and alternative method of promoting the

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degradation of antibiotics.
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Declaration of interests

☒ The authors declare that they have no known competing financial interests or personal relationships
that could have appeared to influence the work reported in this paper.

☐The authors declare the following financial interests/personal relationships which may be considered
as potential competing interests:

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