Following biological treatment, wastewater continues to have endotoxic active materials. However, because there is a trend of potable reuse and because endotoxic active materials potentially have harmful effects on human health, their removal from water is crucial. Lipopolysaccharide endotoxin has hydrophobic groups, and their removal using a coagulation-flocculation alternative is believed to be efficient. Thus, their removal from reclaimed wastewater using the coagulation-flocculation process was assessed. Secondary effluent samples from a wastewater treatment plant located in Sapporo, Japan, were investigated. It was found that this process gave satisfactory results in removing endotoxins, with an optimum removal rate of up to 40.5%. The endotoxin removal was maximized by adjusting the pH at the low range 4 - 5.5, with an aluminum sulfate dose of 80 mg/L. Further increases of the coagulant dose did not improve the removal efficiency. DOC and turbidity removal were at their optimum at higher pH range 5.5 - 6.5. Thus coagulation and flocculation could be considered as the first barrier and should be followed by other treatments to safely reuse reclaimed wastewater.
Potable reuse of reclaimed wastewater is emerging as an alternative to alleviate stresses on conventional freshwater resources and augment potable water supply. Nowadays, we count several potable reuse plants treating wastewater to a potable level. Reliance on potable reuse is expected to grow in coming years. Moreover, groundwater recharge using reclaimed wastewater has become a widespread practice around the globe [
Several other removal techniques such as affinity adsorbents, anionic-exchange chromatography, gel filtration chromatography, and Triton X-114 phase separation among others were employed to remove LPS endotoxin form biological preparations [
Spot samples from the secondary effluent were collected from the settling tank (secondary treatment) of the activated sludge-operated wastewater treatment plant in Sapporo, Japan. The samples were immediately transported to the laboratory, and their characteristics were examined and recorded without delay. The assays were performed in triplicate, and average values are presented.
Jar test experiments were performed to assess turbidity, DOC and LPS endotoxin removal as well as coagulation-flocculation kinetics at various pH values. The secondary effluent water samples were placed in a variable speed ZR4-6 jar test device (SuidoKikoKiasha ltd.), and their pH was adjusted from 4.5 to 8.5 (at an increment of 1) using either 0.1M sodium hydroxide (NaOH) or 0.1M hydrochloridric acid (HCl). According to Pernitsky and Edzwald (2006), favorable pH conditions for alum coagulation generally occur between a pH of 5.8 - 6.5. Then, these samples were subjected to the Coagulation-flocculation (CF) test. Aluminum sulfate, a common coagulant and a preferred reagent with several advantages, including high efficiency at low doses, low cost, low toxicity and ease of availability, was used. The test was performed in a 1-liter beaker at ambient temperature by varying the aluminum sulfate content and pH values (other parameters including rapid mixing speed and mixing time were kept constant). A change of one variable at a time was adopted in this study. First, at a given dose, a wide range of pH values was covered. From this test the optimum pH value for the best LPS endotoxin removal was obtained. The second test involved the variation of coagulant dose while setting the pH at its optimum value obtained from the previous test. As explained earlier, pH was adjusted using sodium hydroxide and hydrochloridric acid. The samples were tested using the following sequence: 60 seconds of rapid mixing at 110 rpm to enhance coagulation, followed by 30 minutes of slow mixing at 25 rpm to favor flocculation, and 30 minutes for settling. These operating conditions were selected randomly. Several coagulation test sets were conducted at different pH values, as mentioned above. The considered operating conditions are summarized in
To assess the treatment efficiency, the samples were analyzed before and after the Coagulation-flocculation process. The water turbidity and DOC were measured in a HACH spectrophotometer, using a HACH kit according to the method 10173 of the HACH water analysis book (HACH, 2001) [
The LAL end point chromogenic assay using a general purpose colorimeter was used to quantify the endotoxins (Anonymous, 2006). A standard curve was established using a negative control of depyrogenated water (Et. and
Parameters | Average value | Standard deviation |
---|---|---|
LPS Endotoxin (EU/ml) | 1490 | 131 |
Dissolved Organic Carbon DOC (mg/L) | 21 | 1.2 |
Turbidity (NTU) | 12.9 | 0.6 |
Electric conductivity EC (μs/cm) | 749 | 69 |
Parameters | Values |
---|---|
Rapid mixing (rpm) | 110 |
Rapid mixing times (seconds) | 60 |
aluminum sulfate doses (mg/L) | 20, 40, 80 and 250 |
pH | 4.5, 5.5, 6.5, 7.5, 8.5 |
beta glucan free) and CSE at endotoxin activities of 1, 0.1, 0.025 and 0.00625 EU/ml. The samples were incubated with the LAL reagent at 37˚C. A general purpose spectrophotometer was used to measure the absorbance at 405 nm. Because absorbance is related to endotoxin activity, the endotoxin activity in the unknown sample was determined by comparison to the standard curve. To validate the readings, positive controls spiked with 0.1 EU/ml of endotoxins were used to determine the recovery ratio. The spike recovery ratio for each sample must be between 50% and 200% to demonstrate a range of insignificant interference and to determine the appropriate sample dilutions. To prevent the pH interference during the LAL assay, TrisHCl buffer was used to maintain the pH at approximately 6.
The total endotoxin activities, DOC and turbidity of water samples, before and after coagulation tests, were measured in triplicates.
An endospecy ES 24 reagent kit for the chromogenic assay of endotoxin was purchased from the SEIKAGAKU Corporation, Japan. The kit consists of a lysate of Limulus Polyphemus (LAL reagent) and a synthetic chromogenic substrate, a buffer solution to dissolve the LAL reagent and depyrogenated water (β-glucan free). The control standard endotoxin (CSE, 90 EU/vial), Pyrocolor Diazo reagent (for use with the endospecy ES 24 kit), and the LAL reagent water were purchased from the Associates of Cape Cod, Inc.
Depyrogenated glass dilution and reaction tubes and depyrogenated pipette tips were purchased from SEIKA- GAKU Corporation, Japan. Other glassware items were washed, rinsed with depyrogenated water, and finally, heat treated for 120 minutes at 250˚C or above.
LPS endotoxin measurements were validated using a spike test. In this test a sample with a known LPS endotoxin is spiked with a known amount of LPS endotoxin. To valid the LAL test the percentage of spike recovered should be between 50% and 200%. This validation indicates that there is no interference (enhancement or inhibition) from test samples. In this study, after the appropriate dilutions, the recovery of endotoxin-spiked samples for the LPS endotoxin detection, using the LAL assay, ranged from 50% to 200%. This validates the LAL assay for the tested samples in this work.
The pH value is an important parameter in determining effective coagulation. Hence, the pH of samples was adjusted from 4.5 to 8.5 (at an increment of 1) then coagulant was added. It is worth mentioning, that the solution pH value decreased in all cases as a function of the aluminum sulfate dose addition.
The effect of the initial pH on endotoxin removal is discussed in this paragraph. The coagulant dose was fixed at
80 mg/L. During the experiments it was observed that the LPS endotoxin, DOC concentrations and turbidity decreased during the laboratory CF test. The average DOC concentration prior to the CF test was equal to 21 mg/l (the average of triplicate measurements). The endotoxin concentration was equal to 1490 EU/ml (average). The average effluent DOC concentrations at the end of the CF test were 11.3 mg/l, 8.9 mg/l, 9.8 mg/l, 10.5 mg/l, and 11.2 mg/l for pH values stabilized at 4.13, 4.63, 5.3, 5.8 and 6.35 after coagulant addition, respectively. As shown in
It is worth mentioning that the percentage of endotoxin removal was at its highest values at lower pH (4 - 5.5). The optimum LPS endotoxin removal was achieved at a pH of 5.3, when 40.6% of the endotoxin was removed (
However, a different trend was observed for the DOC and turbidity removal. The optimum pH for DOC and turbidity removal is 5.8 and 6.35, respectively. This suggests that the main removal mechanism is entrapment as negatively charged species dominates in this range of pH. The turbidity removal ranged from 72.6% to 83.2%, and the DOC removal varied from 45% to 57.3%. The best pH value for the turbidity and DOC endotoxin is in the range of 5.8 to 6.35, while the best pH value for LPS endotoxin removal is rather in the low pH zone (4.3 - 5.3) (
The coagulant dose effect on the endotoxic active material removal is summarized in this paragraph. The samples were flocculated using 20 mg/L, 40 mg/L, 80 mg/L and 250 mg/L aluminum sulfate doses. Since the highest LPS endotoxin removal was observed in the pH range 4 - 5.5, the optimization of aluminum sulfate dosage was performed by adjusting sample pH to 5.5 while varying the alum dosage from 20 - 250 mg/L. As shown in
Comparable shapes of removal percentage curves were observed for DOC and turbidity removal (
It is to be noticed that increasing the aluminum sulfate dosage is not in favor of turbidity removal. In some cases we found that at higher doses of aluminum sulfate (250 mg/L) were often observed to add turbidity to the water (
The process of coagulation/flocculation produced satisfactory results as an alternative for removing endotoxins.
The LPS endotoxin removal using the coagulation flocculation process achieved satisfactory but not excellent results, with up to 40.5% LPS endotoxin removal at its highest efficiency. The LPS endotoxin concentration in the supernatant of the coagulation flocculation process was found to be significantly higher than the values reported for tap water and groundwater (Anderson et al., 2002). Without standards, further treatments are required to meet the current LPS endotoxin levels in tap water and groundwater. Hence, CF is an effective first barrier to reduce endotoxins in the reclaimed wastewater.
In comparison with other treatment alternatives (e.g., sand filtration, membrane filtration, oxidation and UV treatment), coagulation flocculation is less efficient but is still an attractive cheap and affordable method. Indeed, the soil filtration approach reported by Guizani et al. in 2011 exhibited a good removal efficiency (75.6%), but with high instability in time. However, the process is not recommended for the shallow aquifer application. Indeed, long soil columns are required for better efficiency (90 cm). The nano-filtration and reverse osmosis methods have a removal efficiency of more than 90% [
Potable reuse of reclaimed wastewater is becoming a worldwide common trend. Hence efficient removal of potential contaminants found in reclaimed water is of great concern. LPS endotoxins are among the emerging contaminants of concern. In this study, taking advantage of their net negative charge and hydrophobic character, the removal of LPS endotoxin from reclaimed wastewater has been assessed using coagulation flocculation test with the use of aluminum sulfate as coagulant. The coagulation and flocculation process provided satisfactory results in reducing LPS endotoxins in water as a first barrier. The highest endotoxin removal was achieved at an aluminum sulfate dose of 80 mg/L at a pH of 4.69. However, the optimum pH for DOC and turbidity removal was 6.35. The residual LPS endotoxin concentration was significantly higher than the reported value in tap and groundwater. This suggests a need for further treatments for safe potable reuse. Furthermore, even though there is a significant reduction of DOC and turbidity, the remaining turbid matter presents a risk of LPS endotoxin release following the bacteria regrowth. To prevent bacteria regrowth, further treatments are essential. Briefly, the CF process is a satisfactory first barrier for LPS endotoxin removal treatment.
The authors gratefully acknowledge the financial support from the Japan Society for the Promotion of Sciences (JSPS) for the first author, who conducted this research when he was a visiting scholar as an International Research Fellow.
M. Guizani,M. A. Lopez Zavala,N. Funamizu, (2016) Assessment of Endotoxin Removal from Reclaimed Wastewater Using Coagulation-Flocculation. Journal of Water Resource and Protection,08,855-864. doi: 10.4236/jwarp.2016.89070