The use of treatment wetlands plants for protein and cellulose valorization in biorefinery platform.

Different wetland plants were evaluated regarding their potential to be used in further green biorefining platforms to produce soluble protein and cellulose-textile fibers. The results show a higher protein content in the plants grown in treatment wetland conditions, compared with the same species grown in natural conditions, and diverse effect on the content of cellulose, hemicellulose, and lignin, depending on the plant species, more than the growing environment. The TW biomass did not represent a risk regarding accumulation of heavy metals, named Pb, Cd, and Cr, since the studied plants did not present it in their tissues, neither in the roots nor in the leaves. The results regarding cellulose quality of the TW plants showed positive results, having values of molar mass distributions and degrees of polymerization that suggest a suitability to be considered for cellulose-fiber textiles studies. This is one of the first approaches, in the TW field, to establish a new criterion for selecting plant species to be planted in the system, aiming at recovering resources and use them as inputs for biorefineries and sustainable biobased products.

Constructed wetlands and solar-driven disinfection technologies for sustainable wastewater treatment and reclamation in rural India: SWINGS project.

SWINGS was a cooperation project between the European Union and India, aiming at implementing state of the art low-cost technologies for the treatment and reuse of domestic wastewater in rural areas of India. The largest wastewater treatment plant consists of a high-rate anaerobic system, followed by vertical and horizontal subsurface flow constructed wetlands with a treatment area of around 1,900 m2 and a final step consisting of solar-driven anodic oxidation (AO) and ultraviolet (UV) disinfection units allowing direct reuse of the treated water. The implementation and operation of two pilot plants in north (Aligarh Muslim University, AMU) and central India (Indira Gandhi National Tribal University, IGNTU) are shown in this study. The overall performance of AMU pilot plant during the first 7 months of operation showed organic matter removal efficiencies of 87% total suspended solids, 95% 5-day biological oxygen demand (BOD5) and 90% chemical oxygen demand, while Kjeldahl nitrogen removal reached 89%. The UV disinfection unit produces water for irrigation and toilet flushing with pathogenic indicator bacteria well below WHO guidelines. On the other hand, the AO disinfection unit implemented at IGNTU and operated for almost a year has been shown to produce an effluent of sufficient quality to be reused by the local population for agriculture and irrigation.

Microbial communities from different types of natural wastewater treatment systems: vertical and horizontal flow constructed wetlands and biofilters.

The prokaryotic microbial communities (Bacteria and Archaea) of three different systems operating in Denmark for the treatment of domestic wastewater (horizontal flow constructed wetlands (HFCW), vertical flow constructed wetlands (VFCW) and biofilters (BF)) was analysed using endpoint PCR followed by Denaturing Gradient Gel Electrophoresis (DGGE). Further sequencing of the most representative bacterial bands revealed that diverse and distinct bacterial communities were found in each system unit, being γ-Proteobacteria and Bacteroidetes present mainly in all of them, while Firmicutes was observed in HFCW and BF. Members of the Actinobacteria group, although found in HFCW and VFCW, seemed to be more abundant in BF units. Finally, some representatives of α, ß and δ-Proteobacteria, Acidobacteria and Chloroflexi were also retrieved from some samples. On the other hand, a lower archaeal diversity was found in comparison with the bacterial population. Cluster analysis of the DGGE bacterial band patterns showed that community structure was related to the design of the treatment system and the organic matter load, while no clear relation was established between the microbial assemblage and the wastewater influent.

Monitoring the startup of a wet detention pond equipped with sand filters and sorption filters.

The startup of a wet retention pond designed for extended stormwater treatment was monitored by more than one year of continual measurement of hydraulic parameters, nutrients and quality parameters in the pond itself (pH, temperature, dissolved oxygen, turbidity). The data revealed that photosynthesis played an important role for dissolved oxygen and pH for most of the year. Another important observation was that the pond behaved more like a completely mixed reactor than like a plug flow reactor--even though the length to width ratio was as high as 4.5:1. The pond was equipped with sand filters and sorption filters whereby very good nutrient removal efficiencies were achieved.

Pilot-scale comparison of constructed wetlands operated under high hydraulic loading rates and attached biofilm reactors for domestic wastewater treatment.

Four different pilot-scale treatment units were constructed to compare the feasibility of treating domestic wastewater in the City of Heraklio, Crete, Greece: (a) a free water surface (FWS) wetland system, (b) a horizontal subsurface flow (HSF) wetland system, (c) a rotating biological contactor (RBC), and (d) a packed bed filter (PBF). All units operated in parallel at various hydraulic loading rates (HLR) ranging from 50% to 175% of designed operating HLR. The study was conducted during an 8 month period and showed that COD removal efficiency of HSF was comparable (>75%) to that of RBC and PBF, whereas that of the FWS system was only 57%. Average nutrient removal efficiencies for FWS, HSF, RBC and PBF were 6%, 21%, 40% and 43%, respectively for total nitrogen and 21%, 39%, 41% and 42%, respectively for total phosphorus. Removals of total coliforms were lowest in FWS and PBF (1.3 log units) and higher in HSF and RBC (2.3 to 2.6 log units). HSF showed slightly lower but comparable effluent quality to that of RBC and PBF systems, but the construction cost and energy requirements for this system are significantly lower. Overall the final decision for the best non-conventional wastewater treatment system depends on the construction and operation cost, the area demand and the required quality of effluent.

Twenty years experience with constructed wetland systems in Denmark--what did we learn?

Full scale constructed wetland systems for wastewater treatment have been in operation in Denmark since 1983, mainly for the treatment of domestic sewage from small villages. The systems are constructed as soil-based horizontal subsurface flow systems but, because of low soil hydraulic conductivity, surface runoff is evident in most of the systems. Two decades of experience show that soil-based systems are generally efficient in removing suspended solids and BOD, but the removal of nitrogen and phosphorus is lower (typically 30-50%) and the systems do not nitrify ammonium. Contrary to earlier claims, the reeds do not increase the hydraulic conductivity of cohesive soils as much as necessary to secure sub-surface flow. Operation needs of soil-based reed beds are low and normally restricted to emptying of the sedimentation tank, cleaning of the distribution system and mowing of the grass around the system. The dead plant material and accumulated litter on the surface of the systems improve performance after the initial years. A significant number of systems have been shut down or extended with other technologies in order to meet new effluent standards, particularly demands for nitrification. New constructed wetland systems are either compact vertical flow systems which provide good nitrification, willow systems with no discharge or restored wetland systems for nitrate removal. If efficient removal of phosphorus is required, this is achieved by chemical precipitation in the sedimentation tank.

Wastewater treatment in tsunami affected areas of Thailand by constructed wetlands.

The tsunami of December 2004 destroyed infrastructure in many coastal areas in South-East Asia. In January 2005, the Danish Government gave a tsunami relief grant to Thailand to re-establish the wastewater management services in some of the areas affected by the tsunami. This paper describes the systems which have been built at three locations: (a) Baan Pru Teau: A newly-built township for tsunami victims which was constructed with the contribution of the Thai Red Cross. Conventional septic tanks were installed for the treatment of blackwater from each household and its effluent and grey water (40 m3/day) are collected and treated at a 220 m2 subsurface flow constructed wetland. (b) Koh Phi Phi Don island: A wastewater collection system for the main business and hotel area of the island, a pumping station and a pressure pipe to the treatment facility, a multi-stage constructed wetland system and a system for reuse of treated wastewater. The constructed wetland system (capacity 400 m3/day) consists of vertical flow, horizontal subsurface flow, free water surface flow and pond units. Because the treatment plant is surrounded by resorts, restaurants and shops, the constructed wetland systems are designed with terrains as scenic landscaping. (c) Patong: A 5,000 m2 constructed wetland system has been established to treat polluted water from drainage canals which collect overflow from septic tanks and grey water from residential areas. It is envisaged that these three systems will serve as prototype demonstration systems for appropriate wastewater management in Thailand and other tropical countries.

Danish guidelines for small-scale constructed wetland systems for onsite treatment of domestic sewage.

The Danish Ministry of Environment and Energy has passed new legislation that requires the wastewater from single houses and dwellings in rural areas to be treated adequately before discharge into the aquatic environment. Therefore official guidelines for a number of onsite treatment solutions have been produced. These include guidelines for soakaways, biological sand filters, technical systems as well as different types of constructed wetland systems. This paper summarises briefly the guidelines for horizontal flow constructed wetlands, vertical flow constructed wetlands, and willow systems with no outflow and with soil infiltration. There is still a lack of a compact onsite solution that will fulfil the treatment classes demanding 90% removal of phosphorus. Therefore work is presently being carried out to identify simpler and robust P-removal solutions.

Phosphorus removal in constructed wetlands: can suitable alternative media be identified?

Removal of phosphorous in constructed wetlands is limited by the capacity of the media to adsorb, bind or precipitate the incoming P. To enhance P removal and the life span of constructed wetlands the approach might be to use natural sands rich in calcium or iron, to use an alternative 'artificial' medium with high P-binding capacity, or to establish external P-binding filters after the wetland. Our studies focused on the evaluation of calcium-rich materials potentially useful as P-binding media. The materials tested included calcite products, natural sands and seashells. Tests included assessment of physical and chemical properties of the materials, extractions in P-spiked water at different P concentrations to determine P-binding equilibrium isotherms, and column experiments. In addition, full-scale tests were performed with calcite in an external filter. The result showed that equilibrium isotherm is an indicator of the potential P-sorption capacity of the media, although the value is of limited application for the determination of the binding capacity in full-scale systems. The columns showed that the materials do bind phosphorus. However, the binding capacities are still insufficient for the establishment of external P-removal filter; the volumes of the filters would be too large to be of practical use.

Removal of indicator bacteria from municipal wastewater in an experimental two-stage vertical flow constructed wetland system.

The removal of sanitary indicator bacteria (total coliforms, faecal coliforms, and faecal streptococci) was studied in an experimental constructed wetland system consisting of (1) a 2-m3 three-chamber sedimentation tank, (2) a 5 m2 vertical flow constructed wetland, (3) a filter-unit with calcite aimed at removing phosphorus, and (4) a 10 m2 vertical flow constructed wetland. The indicator bacteria were enumerated before and after each unit of the wetland system during four monitoring episodes with different loading conditions. At a hydraulic loading rate of 520-1,370 mm/d, the first-stage vertical flow beds removed about 1.5 log-units of total coliforms, 1.7 log-units of faecal coliforms and 0.8 log-units of faecal streptococci. In the second stage bed receiving lower loadings both in term of concentration and quantity (260-690 mm/day), the eliminations were lower. It was not possible in the present study to identify any seasonal effects, but no measurements were done during summer. Recycling of treated effluent back to the sedimentation tank did not affect elimination. Area-based rate constants for the vertical flow wetland receiving effluent from the sedimentation tank averaged 3.2 m/d for total coliforms, 3.3 m/d for faecal coliforms and 2.1 m/d for faecal streptococci. The rate constants depended on loading rates. It is suggested that filtration is a major removal mechanism for bacterial indicator organisms in vertical flow constructed wetland systems.

Phosphorus removal from municipal wastewater in an experimental two-stage vertical flow constructed wetland system equipped with a calcite filter.

Laboratory studies have indicated that calcite may be used in separate, exchangeable filter units in constructed wetland systems to remove phosphorus. Based on these studies we built a full-scale experimental constructed wetland with a calcite-based filter unit to study its performance, under real-life conditions. The system consists of a 2-m3 sedimentation tank and two vertical flow constructed wetlands. The system has three 0.09 m3 calcite filter-units to study phosphorus removal. The hydraulic loading rate varied between 1.7 and 6.2 m3/d. The residence time in filters ranged from 28 to 99 minutes. Overall the system removed 62 +/- 18% of phosphorus. The removal in the calcite filter was initially good, but after three months all P-filters were saturated. The calcite increased pH by approximately half a unit and released calcium. A total of about 2.2 kg P/m3 calcite was removed by the filter. The first-stage bed receiving effluent from the sedimentation tank consistently removed phosphorus, whereas the second bed sometimes released phosphorus. The first order area-based removal rate constant for total-P in the vertical bed averaged 0.24 +/- 0.20 m/d and was highly dependent on the loading rate. This shows that first order removal kinetics do not satisfactorily describe removal of phosphorus in vertical flow constructed wetland systems with unsaturated flow.

Phosphorus adsorption maximum of sands for use as media in subsurface flow constructed reed beds as measured by the Langmuir isotherm.

The P-adsorption capacities of 13 Danish sands were studied by short-term isotherm batch experiments and related to the physico-chemical characteristics of the sands. The maximum P-adsorption capacities (Q) and P-binding energy constants (b) were calculated using the Langmuir-isotherm model. The Freundlich model was also used, but it was not useful for the description of adsorption phenomena per se since it fitted well P-removal data even if precipitation of P-salts occurred simultaneously. The Langmuir model described the data well (R(2)=0.90-0.99) when precipitation of phosphates did not occur and seems therefore to be useful for describing the adsorption processes per se. The relationships between maximum sorption capacities and physico-chemical characteristics of the sands were investigated using classical univariate and partial least squares regression analyses. Among the physico-chemical properties of the sands, Ca and Mg content, grain size, porosity, bulk density and hydraulic conductivity were significantly related (P<0.1) to the maximum adsorption capacity as estimated by the Langmuir model. Using the maximum P-adsorption capacities, it was estimated how long the P-removal can be sustained with the different sands in subsurface flow constructed reed beds. If the most efficient sand for P-adsorption was used, the adsorption capacity would be used up after about 1 year, while, for the less efficient sands, the P-retention would go on for about 2 months. This suggests that, in order to sustain a long-term P-removal in subsurface flow constructed reed beds, precipitation reactions of insoluble P-salts should be promoted. P-binding energy constants were not significantly related to the physico-chemical properties of the sands, except the Ca content, which showed, however, a low correlation coefficient.

Phosphorus removal by sands for use as media in subsurface flow constructed reed beds.

Sorption of P to the bed sand medium is a major removal mechanism for P in subsurface flow constructed reed beds. Selecting a sand medium with a high P-sorption capacity is therefore important to obtain a sustained P-removal. The objective of this study was to evaluate the P-removal capacities of 13 Danish sands and to relate the removal to their physico-chemical characteristics. The P-removal properties were evaluated in short-term isotherm batch-experiments as well as in 12-week percolation experiments mimicking the P-loading conditions in constructed reed bed systems. The P-removal properties of sands of different geographical origin varied considerably and the suitability of the sands for use as media in constructed reed beds thus differs. The P-removal capacity of some sands would be used up after only a few months in full-scale systems, whereas that of others would persist for a much longer time. The most important characteristic of the sands determining their P-removal capacity was their Ca-content. A high Ca content favours precipitation with P as sparingly soluble calcium phosphates particularly at the slightly alkaline conditions typical of domestic sewage. In situations where the wastewater to be treated is more acid, the contents of Fe and Al may be more important as the precipitation reactions with these ions are favoured at lower pH levels. The maximum P-sorption capacities estimated using the Langmuir-isotherm plots did not correspond to or correlate with the actual amount of P removed in the percolation columns. Hence, the Langmuir-isotherm does not estimate the P-removal capacities of sands. It is suggested that a suitable quick method of screening a selection of potential media for P-removal potential is to perform simple removal isotherm studies using water with a similar chemical composition as the wastewater to be treated. This method will not provide a direct estimate of the P-removal capacity that can be obtained in full-scale systems, but it is a means of comparing the relative performance of potential media.

Removal of nutrients from combined sewer overflows and lake water in a vertical-flow constructed wetland system.

Lake Utterslev is situated in a densely built-up area of Copenhagen, and is heavily eutrophicated from combined sewer overflows. At the same time the lake suffers from lack of water. Therefore, a 5,000 m2 vertical flow wetland system was constructed in 1998 to reduce the phosphorus discharge from combined sewer overflows without reducing the water supply to the lake. During dry periods the constructed wetland is used to remove phosphorus from the lake water. The system is designed as a 90 m diameter circular bed with a bed depth of c. 2 m. The system is isolated from the surroundings by a polyethylene membrane. The bed medium consists of a mixture of gravel and crushed marble, which has a high binding capacity for phosphorus. The bed is located within the natural littoral zone of the lake and is planted with common reed (Phragmites australis). The constructed wetland is intermittently loaded with combined sewer overflow water or lake water and, after percolation through the bed medium, the water is collected in a network of drainage pipes at the bottom of the bed and pumped to the lake. The fully automated loading cycle results in alternating wet and dry periods. During the initial two years of operation, the phosphorus removal for combined sewer overflows has been consistently high (94-99% of inflow concentrations). When loaded with lake water, the phosphorus removal has been high during summer (71-97%) and lower during winter (53-75%) partly because of lower inlet concentrations. Effluent phosphorus concentrations are consistently low (0.03-0.04 mg/L). Ammonium nitrogen is nitrified in the constructed wetland, and total suspended solids and COD are generally reduced to concentrations below 5 mg/L and 25 mg/L, respectively. The study documents that a subsurface flow constructed wetland system can be designed and operated to effectively remove phosphorus and other pollutants from combined sewer overflows and eutrophicated lake water.

Zero-discharge of nutrients and water in a willow dominated constructed wetland.

A novel constructed wetland system has been developed to treat sewage, evaporate water and recycle nutrients from single households at sites where effluent standards are stringent and soil infiltration is not possible. Main attributes of the willow wastewater cleaning facilities are that the systems have zero discharge, the willows evapotranspire the water, and nutrients can be recycled via the willow biomass produced in the system. The willow wastewater cleaning facilities generally consist of c. 1.5 m deep high-density polyethylene-lined basins filled with soil and planted with clones of willow (Salix viminalis L.). The surface area of the systems depends on the amount and quality of the sewage to be treated and the local annual rainfall. For a single household the area needed typically is between 200-300 m2. Settled sewage is dispersed underground into the bed under pressure. When correctly dimensioned, the willow will--on an annual basis--evapotranspire all water from the sewage and rain falling onto the system, and take up all nutrients and heavy metals from the sewage. The stems of the willows are harvested on a regular basis to remove nutrients and heavy metals and to stimulate the growth of the willows. Initial experiences from full-scale systems in Denmark show promising results.

Media selection for sustainable phosphorus removal in subsurface flow constructed wetlands.

Sorption of phosphorus (P) to the bed sand medium is a major removal mechanism for P in subsurface flow constructed wetlands. Selecting a sand medium with a high P-sorption capacity is therefore important to obtain a sustained P-removal. The P-removal capacities of 13 Danish sands were evaluated and related to their physico-chemical characteristics. The P-removal properties of sands of different geographical origin varied considerably and the suitability of the sands for use as media in constructed reed beds thus differs. The P-sorption capacity of some sands would be used up after only a few months in full-scale systems, whereas that of others would subsist for a much longer time. The most important characteristic of the sands determining their P-sorption capacity was their Ca-content. Also the P-binding capacities of various artificial media were tested (light-expanded-clay-aggregates (LECA), crushed marble, diatomaceous earth, vermiculite and calcite). Particularly calcite and crushed marble were found to have high P-binding capacities. It is suggested that mixing one of these materials into the sand or gravel medium can significantly enhance the P-sorption capacity of the bed medium in a subsurface-flow constructed wetland system. It is also possible to construct a separate unit containing one of these artificial media. The media may then be replaced when the P-binding capacity is used up.

Design and performance of the Phyto-Nutri-Tron: a system for controlling the root and shoot environment for whole-plant ecophysiological studies.

A controlled environment system, termed the Phyto-Nutri-Tron (PNT), has been established to study whole plant ecophysiological responses to multiple environmental factors. The PNT is a computer-controlled highly flexible growth facility with independent control of the shoot and the root environment. The facility consists of two growth cabinets each containing four separate hydroponic growth systems. The growth cabinets can be used as assimilation chambers with individual control of temperature, humidity, light, CO2 and monitoring of O2. The hydroponic growth systems are connected to nutrient supply units with disinfection systems and individual control of temperature, pH and oxygen. The ionic composition of the solutions has automated feedback control through a PO4 autoanalyzer and a flow injection analyzer which also analyzes NH4+, NO2- and NO3-. Other ions are automatically monitored by ICP-AES. The system has automated calibration procedures of the analytical equipment and prolonged studies of plant growth can be performed under constant environmental conditions. This paper describes the design and construction of the PNT, the results of a number of tests showing the degree of control of environmental factors and the results of a comparative study on NH4+ and NO3- uptake kinetics by Juncus effusus conducted in the PNT demonstrate the use of the PNT in ecophysiological studies.

Modulation of cytosolic sexual steroid receptors in autochthonous methylnitrosourea-induced rat mammary carcinoma following application of 2-chloroethylnitrosocarbamoyl-L-alanine linked to oestradiol or dihydrotestosterone.

This study concentrated on the influence of 2-chloroethylnitrosocarbamoyl-L-alanine (CNC-L-ala) linked to oestradiol (CNA-L-ala-E2) or dihydrotestosterone (CNC-L-ala-DHT) in position 17 of the respective steroid hormone on tumour growth and receptor kinetics of methylnitrosourea-induced rat mammary carcinoma. Both compounds almost completely arrested logarithmically growing mammary carcinoma of Sprague-Dawley rats: in the first week CNC-L-ala-E2 blocked the growth of these tumours by 92% compared to untreated control animals while, in animals treated with the physically equimolar mixture of CNC-L-ala and oestradiol (positive control), tumour growth was inhibited by 51% only. CNC-L-ala-DHT arrested the tumour growth in the first week by 95%, while the respective positive control (CNC-L-ala plus dihydrotestosterone) effected a growth inhibition of 71% compared to the untreated control. These results correlate well with the influence of both drugs on the cytosolic receptor content of sexual steroid hormones in the tumours. CNC-L-ala-E2 depleted the content of oestradiol receptors and kept it down for a week, while concomitantly the content of progesterone receptors increased considerably and that of androgen receptors showed a short-lived decrease. CNC-L-ala-DHT depleted androgen receptors as well as progesterone receptors. The content of androgen receptors remained low for a week, while that of progesterone receptors recovered within 8 days. The content of oestrogen receptors showed a moderate decrease.

Androgen-linked alkylating agents: biological activity in methylnitrosourea-induced rat mammary carcinoma.

This article gives a comprehensive survey on the anticancer activity of nitrosoureas linked to steroidal androgens in methylnitrosourea (MMU)-induced rat mammary carcinoma. cis-Androsterone, testosterone, 19-nortestosterone and 5-alpha-dihydrotestosterone were used as carrier hormones and were linked to various cytotoxic N-[N'-(2-chloroethyl)-N'-nitrosocarbamoyl] (CNC)-aminoacids and to N-(2-hydroxyethyl)-N'-(2-chloroethyl)-N'-nitrosourea hemisuccinate (HECNU-hemisuccinate). In the MNU-model used esters of dihydrotestosterone (DHT) invariably were more active and less toxic than those of testosterone, nortestosterone and cis-androsterone. Within the DHT esters of CNC-aminoacids those of CNC-glycine, CNC-methionine and CNC-alanine showed the highest antineoplastic activities and superiority compared with equimolar dosages of their unlinked mixtures. Additionally, CNC-alanine-DHT ester had the highest therapeutic ratio of all agents investigated. HECNU-hemisuccinate-DHT ester, on the other hand, achieved even higher antitumor activity at the optimal dose but had a narrower therapeutic ratio. No obvious correlation between antineoplastic efficacy and receptor binding affinity could be demonstrated, but, to be active, a conjugate apparently had to have some receptor binding affinity for both androgen and progesterone receptors. The results obtained indicate that linking antineoplastic agents to transport molecules with affinity to steroid receptors is a highly promising approach to obtain drugs with specific activity in steroid receptor containing tumors.

Nitrosoureas. Modes of action and perspectives in the use of hormone receptor affinity carrier molecules.

Mechanisms of DNA adduct formation by antineoplastic 2-chloroethyl-N-nitrosoureas (CNUs) and of DNA damage induced by these compounds are discussed. CNUs are alkylating agents that form DNA-DNA cross-links as well as 2-chloroethylated and 2-hydroxyethylated adducts, the N-7-position of guanine being the predominantly alkylated site. A close correlation exists between the potential of a given compound to induce DNA-DNA cross-links and its antineoplastic effectiveness. However, levels of DNA-DNA cross-linking in bone marrow and extent of myelosuppression as measured in rodents are also closely correlated. The design of new cross-linking analogues capable of directing the antineoplastically relevant activity predominantly to the target tumour appears therefore to be of great promise. Cross-linking agents have been attached to a variety of steroid hormone carrier molecules and the conjugates have been tested in structure-activity studies using hormone-receptor containing animal tumours. These studies have revealed that some hormone-linked antineoplastic agents are highly effective in receptor positive experimental tumours and are superior to mixtures of unlinked alkylating agents with hormones. Indications for a relative enrichment of DNA damaging effects in the tumour tissue and for reduced myelotoxicity have been obtained with specific hormone conjugates.