NTA biodegradation and removal in subsurface sandy soil

Wat. Res. Vol. 20, No. 3, pp. 345-349, 1986 Printed in Great Britain. All rights reserved

0043-1354/86 $3.00+ 0.00 Copyright © 1986 Pergamon Press Ltd

NTA BIODEGRADATION A N D REMOVAL IN SUBSURFACE SANDY SOIL NAM H. B ~ I ( and NICHOLAS L. CLESCERI Department of Chemical Engineering and Environmental Engineering, Rensselaer Polytechnic Institute, Troy, NY 12181, U.S.A.

(Received August 1985) Abstract--This lab-scale study examined the biodegradation and removal of nitrilotriacetic acid (NTA) in the subsurface environment, mainly sandy soils. Batch tests indicated that NTA adsorption on the sandy soils played a minor role in its removal in these soils. Removal of NTA was investigated in 50.5 mm i.d. by 1.17m long soil columns under unsaturated conditions at 15°C. Septic tank effluent containing 20 mg NTA 1-1 was dosed to soil columns four times a day at an overall loading rate of 1 gpd ft -2 for a 43-day period. This feed NTA concentration was routinely reduced to a steady-state concentration of 0.1 mg 1-1 by passage through the 1.17 m of soil, after an indigenous soil microflora became sufficiently established over a 25 day period. In addition, the results of samples taken on day 21 demonstrated that greater than 75% removal of NTA can be expected in a soil depth of less than one-third meter.

Key words--nitrilotriacetic acid (NTA), subsurface sandy soil, biodegradation, adsorption

INTRODUCTION Nitrilotriacetic acid (NTA) has been considered as a substitute for phosphate in detergents because of its chelating properties. If N T A is to replace condensed phosphates in detergents throughout the United States, a large a m o u n t o f this substance could be introduced to the environment. Therefore, it is important to understand N T A removal in the subsurface environment and the potential for its entry into groundwater created by widespread use in areas where the septic tank/tile field system is the preferred method of liquid waste disposal. A number of previous studies have shown degradation o f N T A under a variety of conditions, i.e. complete degradation of N T A in nine different soils under aerobic conditions (Tiedje and Mason, 1974), complete degradation through unsaturated soils and slow degradation in saturated groundwater (Dunlap et al., 1972), a fairly rapid rate of biodegradation in oligotrophic groundwater (Larson and Ventullo, 1983) and complete removal of N T A during soil percolation (Hrubec and van Delft, 1981). Klein (1971) concluded that under normal conditions N T A removal in septic tank/tile field systems was virtually complete, except in a seriously ponded condition with no dissolved oxygen, when the removal was only 10%. Despite a limited number of earlier references on N T A removal in soil, the need for examining the fate of N T A in soil located between septic tank/tile field systems and a shallow groundwater table was perceived. During percolation through soil systems, N T A from detergents could be subject to sorption and microbial degradation. This study examined 345

these removal mechanisms in New Y o r k soils and consisted o f two phases, i.e. sorption studies and column biodegradation studies, each of which was performed utilizing an upstate New Y o r k soil and a Long Island soil. The focus of this study was to attempt to demonstrate unequivocally the ultimate removal of N T A in these soils. The experimentation included semicontinuous-flow soil column studies, in which the ultimate degradation of N T A could be explored. This was accomplished by measuring 14CO2 evolution not only in the column effluent but also in the column headspace, utilizing uniformly labelled [14C]NTA. MATERIALS AND

METHODS

The experiment was designed to investigate NTA removal by adsorption and biodegradation in a subsurface soil environment, particularly in properly operating tile field systems. Uniformly labelled [14C]NTA and a liquid scintillation technique were utilized in this study as an accurate and simple measurement of NTA.

Chemicals Uniformly labelled [t4C]NTA was obtained from Procter & Gamble Co. (Cincinnati, Ohio). This t4C-labelled NTA solution, having a concentration of 1 mg ml-l and a specific activity of 42 #Ci mg -I (95% purity based on HPLC), was stored in a freezer. An unlabelled stock solution of nitrilotriacetic acid, trisodium salt monohydrate (NTA-NA3) (Aldrich Chemical Co., Inc.) was prepared at a concentration of 1 g 1-~ with distilled water. For the adsorption study, the NTA-Na 3 solution was stoichiometrically mixed with [14C]NTA to produce 2, 6 and 20 mg NTA 1- l stock solutions, each with a specific activity of 31.5 nCi ml-I. All three stock solutions were sterilized by 0.45gm membrane filtration (GN-6, Gelman Scientific, Inc., Ann Arbor, Mich.), using a preautoclaved filtering set. For the column study, the stock solution was prepared by spiking NTA-Na 3 with [14C]NTA, yielding a specific radio-

346

NAM H. BAEK and NICHOLAS L. CLESCER1

activity of 0 . 6 3 # C i m l ~ and a N T A concentration of 0.44 mg ml ~. Filtration through 0.45 # m membranes for sterilization was also performed with subsequent storage at 4cC.

Septic tank samples Effluent wastewater used in the biodegradation studies was obtained from a septic tank serving an office building. This effluent contained approx. 1 3 0 m g l -t total organic carbon (TOC), 1 0 0 m g l t biochemical oxygen demand (BOD), 6.0 mg P 1-~ total phosphorus and 0.4 mg N lnitrate-nitrogen. The total bacteria count (Acridine Orange Method) of this sample was 4.4 × 106ml % All of these analyses were conducted using the procedures in Standard Methods (APHA, 1980). The septic tank effluent was collected twice during this study and stored at 4°C. Weekly measurement of its total organic content indicated no degradation occurred at 4°C. Soils Soil for this study was obtained from two sites in New York State. A Long Island soil was obtained from the South Shore area of Suffolk County near Center Moriches in a residential area and collected by auger between the depths of 0.6-2.4 m below the surface. The second soil, which had a sandy texture similar to the Long Island soil, was collected from an upstate location (Bolton Landing in Warren County). Both soils were virgin in nature and were characterized as shown in Table 1. Sorption studies The soil was oven-dried at 105°C overnight and sieved with a 2 m m screen. A 50 g sample of the soil was placed into a 125 ml Erlenmeyer flask and autoclaved at 121°C for l h (Klein, 1971). A 7 0 m l sample of a sterilized N T A solution (2, 6 or 20 m g l -I) was added to a soil flask in triplicates. The flasks were shaken continuously at 200 rpm (New Brunswick Scientific Co., Model G-2) at 10°C, an annual average subsoil temperature in New York state. Aseptic techniques in Standard Methods, Part 900 (APHA, 1980) were employed to prevent possible microbial degradation of N T A during the absorption studies. At each N T A concentration, triplicate blanks (containing no soil) were used to determine whether N T A loss other than by adsorption occurred. A standard agar test in Standard Methods, Part 902A (APHA. 1980) confirmed no bacterial contamination in the test flasks during the sorption studies. On day 3, a 5 ml sample was taken from each flask and centrifuged to separate the liquid from the soil. A 1 ml aliquot of the supernatant was added to 10 ml of liquid scintillation cocktail solution (Scintiverse, Fisher Scientific Co.) and then analysed for ~4C by Liquid Scintillation Spectrometry (Model SC-30, Intertechnique Instruments, Inc.). Samples were also taken on day 6 to assure that equilibrium in the adsorption process had been attained. Quantities of N T A adsorption by the soils were calculated from subtraction of the equilibrium concentrations from the initial concentrations as follows: O = --C-'Z (-) × 1/~Ci 103 ,ug n 2.22 x 106dpm x 42/~Ci x ?' x 70ml

q = Q/M where Cj = final counting rate (cpm), C, = initial counting rate (cpm), M = total a m o u n t of soil used for adsorption (50 g, dry wt), Q = total a m o u n t of N T A adsorbed in a flask (/~g), q = adsorptive capacity (/2g g - l ), y = mixing ratio of total N T A to radioactive NTA, = counting efficiency (0.9 cpm/dpm).

Table t. Soil analysis Characteristics Soil texture Moisture content Effective diameter Coefficient of uniformity pH Cation exchange capacity Total organic carbon-C Total phosphorus-P Total inorganic nitrogen-N

Long Island soil

Bolton Landing soil

Sandy 3.6% 0.188 mm 2.35 6.25 1.66 mequiv/100 g 50 mg kg ~ 85 mgkg ' 7.54mgkg ~

Sandy 6.3% 0.074 mm 6.29 7.85 4.45 mequiv/100 g 160 mg kg t 1200mgkg ' 9.06mgkg

Column biodegradation study The nonsterilized soils from which stones, roots and other debris had been removed were packed into 50.8 m m i.d. by 1.8 m long glass columns to produce soil columns which were 1.17 m long. The columns had a porosity of 25% as estimated by the volume of water required to saturate the soil columns. A standard percolation test was conducted to determine the allowable loading rate of this soil system in compliance with Manual of Septic T a n k Practices (U.S. PHS, 1967). Duplicate columns of each soil were used. Each column was composed of three 0.6 m tubes and two intermediate sampling ports between each tube (Fig. I). The second and third tubes consisted of 3.8 cm pea gravel, 48.3 cm soil and 3.8 cm pea gravel in series. The first tube was specifically designed to simulate the introduction of septic tank effluent into a tile field, i.e. subsurface application and air penetration via head space purging as shown in Fig. 1. The percolation rate of 16-20 min in J, which was obtained in the standard percolation test, allowed a loading rate of 1 gpd ft -2 (U.S. PHS, 1967). For the biodegradation experiments, the columns were dosed four times per day for 43 days at an application rate of 88 ml day -t (1 gpd ft -2) for each column by means of a peristaltic p u m p operating for l0 min at each dosing time. The columns were maintained at a temperature of 15+C, controllable by jacketed coolant circulation. At each application, a mixture of 21 ml of septic tank effluent and a 1 ml of stock NTA solution (0.44 m g N T A ml L) was introduced to the mid-gravel layer of the first tube resulting in a concentration of 20 mg N T A 1- ~. This feed concentration would be encountered in tile fields when N T A containing detergent is used in household laundering. As shown in Fig. 1, air was passed continuously over the head space of each column to mitigate against any oxygen limitation, and to purge carbon dioxide released from biological activity in the soil into two sequential traps (Top CO 2 Traps) containing 40 ml of 5 N potassium hydroxide. After passing through the top CO 2 traps, air was then purged into a saturated Ba(OH)2 solution for a visual check on any CO 2 loss. The percolate from each column was collected in a 500 ml filtering flask which contained 10ml of 1 N potassium hydroxide solution which prevented any biological activity during the 2-day collection period. The outlet of each percolate collection flask was attached to two 100ml test tube traps (Bottom CO 2 Traps) in series containing 50 ml of 5 N potassium hydroxide solution. The four percolate collection flasks and their respective CO 2 traps were removed from the columns every 2 days. The percolate collected from the bottom of each column was acidified to pH = 2 with 5 ml of I0 N hydrochloric acid, and then purged with nitrogen gas for 10min to release any carbon dioxide into the preattached bottom CO 2 traps. During N 2 purging, the lack of any CO 2 loss was visually confirmed by passing the outlet gas through a saturated Ba(OH)2 solution. Following this nitrogen purging, 1 ml aliquots from each of the four percolate collection flasks and each of the

347

NTA biodegradation and removal

~

~ Air

1

Vent

COz Traps

Bo(OH) z

Headspace

Sampling port

Pump

Column influent of

septic tank effluent plus NTA

E ()

Percolate /~ collection / flask ( COz Traps

Fig. 1. Flow diagram of the NTA column study. bottom CO2 traps were injected into I0 ml of liquid scintillation cocktail solution and analyzed for 14C by Liquid Scintillation Spectrometry. The non-purgable radioactivity in the percolate collection flask was calculated as NTA. The same sampling procedure and analysis were applied to the top CO2 traps without N2 purging. Due to the time constraints, intermediate point sampling in the four columns was conducted only once, using the same technique involved in bottom percolate analysis. The counting analysis was calibrated with a standard [14C]Toluene(4.5 x 105dpm ml- ~, New England Nuclear). The overall counting efficiencies were found to be 86% for [14C]NTAanalysis and 83% for [14C]CO2 analysis.

RESULTS AND DISCUSSION

Sorption studies Adsorption by both soils reached an equilibrium state after 3 days. This was verified by observing that the samples at day 6 showed no sign of further adsorption on the soils. Therefore, the experimental results of the samples taken on day 3 were used to calculate the adsorptive capacity of the soils. The results are presented using the averages of the triplicate samples. The Long Island soil possessed adsorptive capacities of 1.64, 3.93 and 7 . 5 6 # g g -~ at 0.8, 3.2 and 14.6mgNTA1-1, respectively, adsorptive capacities of the Bolton Landing soil were 0.39, 1.03 and 3.34#gg-~ at 1.7, 5.3 and 17.6mgNTA1-1, respectively. The results of the NTA sorption studies with the Long Island soil and

the Bolton Landing soil can be best described by a Freundlich isotherm, i.e. when these data were plotted on log-log scale, a straight-line relationship was obtained. During the 3-day equilibrium period, there was no evidence of NTA degradation, determined by the fact that ~4C-radioactivity of the liquid in the blank flasks remained constant and no evidence of microbial growth in the agar tests was observed. The Freundlich isotherms shown in Fig. 2 indicate that the amount of NTA adsorbexl on the soils was proportional to its concentration in the liquid phase. The adsorptive capacity of the Long Island soil is greater than that of the Bolton landing soil over the range of concentration tested in this study. It is predictable that the Long Island soil, containing low organic content (TOC = 0.5%), may possess more 1o~>" '~

Long Island soil

I

~.z

+

o o

/ O.1 0.1 Equilibrium

I 1

Bolton Landing soil I 10

concentration

I 100 (mgNTA

1-1 )

Fig. 2. Freundlich isotherms of NTA adsorption on the soils at 10°C. Data represent the averages of the triplicates.

348

NAM H. BAEK a n d NICHOLAS L. CLESCERI J-- 2.0

90

- ....

d E

Long Island soil Bolton Landing soil

o>

8 ~ z

95

1.O

or*

"o n,- 0.0 0

100 10

20

30

40

Operation time (days)

Fig. 3. Residual NTA concentrations in percolates after passing through a soil depth of 1.17m at 15°C. Feed concentration was 20 mg NTA 1- ~ and data represent the averages of soil duplicates. adsorptive sites for hydrophilic compounds than the Bolton Landing soil (TOC = 1.6%). These results agree with the conclusion (Hassett and Means, 1980) that the hydrophobic nature of soils with a measurable organic content reduces the sorption of water soluble compounds, such as NTA. Once all adsorptive sites of the soils are occupied, no further adsorption of NTA on the soils is expected unless these sites are regenerated by other processes, such as desorption, chemical reaction and/or bigdegradation. Therefore, adsorption on these soils itself cannot be considered as an ultimate removal process for NTA. Sorption, however, provides some retardation of NTA movement during percolation through the soils.

Column biodegradation studies The NTA concentrations in the percolates over the course of the 43 day test period are presented using the averages of the soil duplicates (Fig. 3). NTA concentrations in the percolates for the first few days (day 0-day 5) were essentially undetectable, undoubtedly attributable to retardation by adsorption on the soils. As the experiment progressed, NTA concentrations in the percolates reached as high as 1.3 mg I-L Following this peak concentration occurring on about day 15, a continuous decline in the percolate NTA concentrations to a steady state concentration of 0.1 mg 1-] was observed in all columns. This may be interpreted as a 15 day period being required to establish a sufficient microbial population in the soils. To support this inference further, a

constant increase of 14CO2 recovery in the CO 2 traps after day 15 was observed as shown by the nearly constant slopes of the curves in Fig. 4. It is, however, interesting to note that the removal of NTA was still greater than 90% for the transitory period (day 1-day 25). It is apparent that during the initial 25 days these soil systems became stabilized to degrade NTA, indicated also by the attainment of nearly steady state conditions for zaco 2 recovery after day 25 (Fig. 4). Consequently, the percolate NTA concentrations decreased to a steady-state level of about 0.1 mg 1-~ for the remainder of the experiment. Before any inorganic carbon-14 from NTA degradation is measurable, an initial lag of 4-7 days was observed (Fig. 4), which coincides with the retardation of NTA movement by adsorption during this period. Thompson and Duthie (1968) also observed a lag in NTA degradation when compared to glucose degradation. As seen in Fig. 4, the rate of inorganic carbon-14 recovery (expressed as the sum of 14CO2 and 14CO3) slowly increases and eventually approaches a steady-state level of 60-70% recovery. The remainder of this carbon (30-40%) is attributable to substances remaining in a column, such as inorganic carbon, NTA and biomass. In order to understand the rate of NTA degradation in the soil columns as a function of soil

2O i L o n g i s l a n d soil

100

v >,

8_

--

X

Boiton L a n d i n g soil

~ lO Long Island soil

~ 5o .÷" s+"

~'x

Ol ~ ' + ~ + " I I 10 20 30 Operation time (days)

I 40

Fig. 4. The recovery of inorganic carbon- 14 (14CO2+ 14CO3) in the CO2 traps from NTA degradation. Data represent the averages of soil duplicates.

0

30

60 S o i l depth

Fig.

5. N T A

concentrations

of

90

120

(cm)

percolates

taken

by

intermediate sampling at a given soil depth on the 21st day of operation. Data represent the averages of soil duplicates.

NTA biodegradation and removal column length, samples were taken at day 21 from two intermediate sampling ports in each column. Figure 5 demonstrates that NTA removal occurred readily with a resultant degradation of 76-98% at a depth of 20.3 cm in all columns, corresponding to a liquid detention time of approx. 3 h. Hrubec and van Delft (1981) also found even faster NTA disappearance with depth in their study of artificial groundwater recharge, i.e. NTA was not detectable at a depth of 10cm below the bed surface (approximated detention time = 1 h). Overall, the mechanism responsible for the majority of the NTA removal observed in these soils was biodegradation. Also, most of the NTA degradation occurred in a relatively short passage through the soils, once the microbial populations became sufficiently established.

CONCLUSIONS Results of this study demonstrate that: (1) Adsorption played a minor role in NTA removal in the test soils (Long Island soil and upstate New York soil). (2) Concentrations of NTA in the applied septic tank effluent (20 mg 1-~) were reduced to a steadystate concentration of 0.1 mg !-1 ( > 9 9 % removal), percolating through 1.17 m of both the Long Island soil and the Bolton Landing soil, after a period of 25 days. (3) An initial time period of 25 days elapsed before the soil columns attained maximum removal of NTA, undoubtedly a result of the time required for microbial populations to become sufficiently established.

349

However, NTA removal was still >90% during this transitory period. Acknowledgements--This research was supported by grants

from The Procter & Gamble Co. The authors wish to thank Dr Lenore S. Clesceri of the Department of Biology, Rensselear Polytechnic Institute for helpful discussions and suggestions during this study. REFERENCES

APHA (1980) Standard Methods for the Examination o f Water and Wastewater, 15th edition. American Public Health Association, Washington, D.C. Dunlap W. J., Cosby R. L., McNabb J. F., Bledsoe B. E. and Scalf M. R. (1972) Probable impact of NTA on groundwater. Ground Water 10, 107-117. Fuller W. H., Korte E. E., Niebla E. E. and Alesii B. A. (I 976) Contribution of the soil to the migration of certain common and trace elements. Soil Sci. 122, 223-235. Hassett J. J. and Means J. C. (1980) Sorption properties of sediments and energy-related pollutants. EPA-600/3-80041, EPA Athens, Ga. Hrubec J. and van Delft W. (1981) Behaviour of nitrilotriacetic acid during groundwater recharge. Water Res. 15, 121-128. Klein S. A. (1971) The fate of NTA in septic-tank and oxidation-pond systems. SERL report No. 71-4, Sanitary Engineering Research Laboratory, University of California, Berkeley, Calif. Larson R. J. and Ventullo R. M. (1983) Biodegradation potential of ground-water bacteria. Proceedings o f the Third National Symposium on Aquifer Restoration and Ground-Water Monitoring, pp. 402-409. Worthington,

Ohio. Thompson J. E. and Duthie J. R. (1968) The biodegradability and treatability of NTA. J. Wat. Pollut. Control Fed. 40, 306-319. Tiedje J. M. and Mason B. B. (1974) Biodegradation of nitrilotriacetate (NTA) in soils. Soil Sci. Soc. Am. Proc. 38, 278-283. U.S. Public Health Service (1967) Manual o f Septic Tank Practices. Public Health Service Pub. 526. U.S. Government Printing Office, Washington, D.C.