WO2002042228A1

WO2002042228A1 - Bioadsoprtion process for the removal of colour from textile effluent

Info

Publication number: WO2002042228A1
Application number: PCT/GB2001/005167
Authority: WO
Inventors: Majeda Khraisheh; Margaret Conlon
Original assignee: Questor Technologies Limited
Priority date: 2000-11-25
Filing date: 2001-11-23
Publication date: 2002-05-30
Also published as: AU2002220820A1; GB0028814D0

Abstract

A process for the bioadsorption of a dye using a bioadsorbent material is described. The bioadsorbent material is a dye-degrading bacterium on or in an absorbent support material. Preferably the bacteria in Shewanella Putrefacrens, and the dye is an azo-dye. A number of absorbent materials can be used as a support material for the present invention. Materials other than carbon may also be used in this process, including materials known to be able to support a biofilm, which are used in water treatment plants to degrade the sludge. With the colour removal process the support material takes on a much more significent role within the process i.e. that of adsorption of the dyes, mainly reactive dyes. It has been found that the microbial activity of the bacterum is significantly promoted by its support on activited carbon, preferably granular activited carbon (GAC). Moreover, such activity does not diminich the primary function of the carbon which remains that of an adsorbent.

Description

BIOADSORPTION PROCESS FOR THE REMOVAL OF COLOUR FROM TEXTILE EFFLUENT

The present invention relates to a process for bioadsorption for removal of colour from textile effluent.

Within the textile industry there is a pressing need to develop a cost-effective process for colour removal which is capable of meeting recently imposed environmental consent levels. Reactive dyes in dye- house effluents discharged to sewers persist after passage through a sewage treatment works, causing subsequent colouration of waterways. Further complications arise from the large number of components, which make up the textile waste and from a lack of homogeneity as a consequence of changing production operations.

A range of end-of-pipe treatment options across different scientific disciplines has been suggested for colour removal, however disadvantages associated with each technique and the reluctance of the industry to adapt unproven laboratory technologies to full-scale have prevented their application. Suppliers, often specialising in one area, have driven the market in recommending single treatment solutions despite indications from both literature and experience that a single method will not provide a cost-effective solution. Hence, colour removal remains a major problem for the textile industry.

One example is the degradation of azo compounds, of which reactive dyes are an example, to aromatic amine under aerobic and anaerobic conditions. All azo dyes are characterised by one or more azo (- =N-) linkages. The colour of azo dyes is due to the azo bond associated chromophores . Little information ^* about the structure of recently used dyes is available, but it is known that the reactive dyes, under alkaline conditions, produce a reactive vinyl sulfone group. This usually enables the dyes to bond to fabrics through nucleophilic addition. Competing with this reaction, at the elevated temperatures associated with dyeing processes, is a hydrolysis reaction.

These classes of azo dyes include mainly acid dyes. However, hydrolysed reactive azo dyes, containing carboxyl or sulfonyl groups degrade poorly due to their high solubility, and degradation can be inhibited by increasing concentration of the dye. Moreover the hydroxyl group of the dye- akes it more polar and hydrophilic than the vinyl sulfone form, preventing it from being adsorbed. In anaerobic sludge digesters, the reductive cleavage of the azo bond is only a partial degradation step, converting azo compounds to aromatic amines. Through this step may remove the colour of the .dyes, it does not effectively eliminate the chemical from the wastewater. Hence, anaerobic degradation in conventional terms cannot be used as a final step in the treatment of hydrolysed azo dyes.

In general, sewage treatment works have found it difficult to remove colour from the wastewater received, largely due to high levels of reactive dyes in the effluent. Where the textile companies producing this colour have been unable to reduce the levels of colour discharged, the local authority are forced to upgrade their treatment works by the inclusion of tertiary treatments, purely to de-colour the effluent prior to discharge to surface waters. In these instances, most notably in the Severn Trent area, the cost to the producers has been in the order of £0.3 to £0.5/m³ discharged. For a small to medium sized textile dyer, this has seen increases in trade effluent charges of up to £50k .per year.

According to one aspect of the present invention, there is provided a process for bioadsorption of a dye, particularly an azo-dye, by a bioadsorbent material comprising a dye-degrading bacterium on or in an absorbent support material. The dyes are usually colour dyes such as azo-dyes, including reactive dyes.

A number of bacteria are known to utilise the azo aromatic structures of other dyes. One class is reported to have an ability to degrade certain classes of dyes. These include Bacillus Gordonae al . , Ba cill us Beneovorans al . , Pseudomonas Putida and Pseudomonas sp. , and some known metal reducing bacteria.

The bacteria can be located on or in the support material in any known fashion, e.g. by biofilm growth of the bacteria on the material.

A number of absorbent materials can be used as a support material for the present invention. For example, EA207, C207, Centaur, Chilean lignite carbon and lignite coke carbon. Carbons classified as H (carbons which form an alkaline suspension and show a relatively high H⁺ adsorption capacity compared to OH^" ions) may be preferred. Materials other . han carbon may also be used in this process, including materials known to be able to support a biofilm, which are used in water treatment plants to degrade the sludge. With the colour removal process the support material takes on a much more .signi icant role within the process i.e. that of adsorption of the dyes, mainly reactive dyes. A carbonate of calcium material from marine origin can also be used as an alternative support material for bacterial cells. The product obtained from De Jong eco Services in the Netherlands is called Algo- filter. The real name is Lithothame Calcaneum. According to the manufacturers, the specific surface area for l-2mm pellets is around 5m²g^-1. Some of the current uses of this material is in the absorbance of chemical spills, support for pre-selected micro- orgamis s used in sludge treatments and in water filtration.

The bacterium Shewanella Putrefacrens is disclosed in UK Patent Application No 2316684 of British Textile and Technology Group. This document is incorporated herein by way of reference. This bacterium has demonstrated an ability to effectively degrade reactive dyes such as azo dyes by utilising such dyes as terminal electronic receptors.

It has been found that the microbial activity of this bacterium is significantly promoted by its support on activated carbon, preferably granular activated carbon fGAC) . Moreover, such activity does not diminish the primary function of the carbon which remains that of an adsorbent.

Some specifications and grades of suitable activated carbons are listed below. Carbon [1) Origin Surface Bulk Iodine value area density (mg/g) (m²/g) (g/cm³)

F-400 Bituminou 1100 0. ,43 1050 s coal

Centaur Bituminou 767 -- 800 s coal

EA207 Coal 950 0. .50 900 C207 Coconut 1050 0. .49 1000 shell

Coke lignite 300 0. ,45 -- carbon Chilean Lignite Lignite

The particular combination of the bacterium Shewanella Putrefacrens and activated carbon provides a means of achieving an economic design capable of being scaled up to the required industrial level.

Thus, according to a second aspect of the present invention, there is provided apparatus for the bioadsorption of dyes, which apparatus allows for the passage of dyes across bioadsorbent material comprising a dye-degrading bacterium such as Shewanella Putrefacrens, and an adsorbent support material such as activated carbon.

Preferably, the apparatus is a multistage process, having a continuous aerobic reactor followed by one or more columns in a fixed bed configuration. The multi-stage process should be designed to meet the requirement of bacterial growth as well as the strict conditions required upon which the microbial film degrades colour.

The apparatus can be of any suitable size, e.g. a stand alone industrial plant, or a cartridge for use in a larger system or unit. The cartridge could be a replaceable fitting.

Preferably, the process of the present invention involves treating dyes including reactive dyes, with a relatively short hydraulic retention time, whilst achieving relatively long solid retention times-. Obtaining biomass retention significantly reduces the volume of the reactor or other vessel required. Moreover, it offers a great advantage over contact processes for dealing with dyes, Including reactive dyes, by reducing the problems associated with solid/liquid separation.

Preferably, the process of the present invention is a continuous fixed-bed bioadsorption process.

According to another aspect of the present invention, there is provided a method for bioadsorption of a dye comprising passing the dye across a bioadsorbent material such as Shewanella Putrefacrens on or in an absorbent support material such as activated carbon. According to a fourth aspect of the present invention, there is provided use of a bioadsorbent material such as Shewanella Putrefacrens in the bioadsorption of dyes such as reactive dyes.

An example of the invention is now generally described by way of example only and with reference to the accompanying graphs and drawings Figures 1 to 8.

Experimental

Culture Growth The Bacteria strain used in this work was supplied by British Textile Technology Group in ampoules of freeze dried culture, which was reconstituted using a sterile nutrient broth and subsequently onto sterile plates of nutrient agar. The details of the experimental techniques used for bacterial growth, media preparation and plating are given in GB2316684A. These samples were incubated at the appropriate temperature, 30-35°C and stored under refrigerated conditions and re-plated every six weeks. The culture was checked routinely under the microscope for contamination. Culture media for subsequent experiments were inoculated from these initial cultures. Conical flasks containing 50-100 ml of sterile liquid soya broth media were inoculated from the solid media cultures and incubated in a shaker/ incubator at a shaker speed of 180 rpm, to maintain dissolved oxygen levels, and at appropriate temperature. Samples taken from the small shake flasks were then used to inoculate the aerobic reactor used in the bioadsorption system. The following Table 1 gives a summary of the optimum growth conditions for this bacteria.

Spectrophotometric Analysis

During the microbial degradation of the dyes, it has been assumed that the amount of degradation is directly proportional to the reduction in dye colour. It is likely that the bacteria breaks down the azo bond in the dye structures, but would not oxidise the dyes completely into carbon dioxide and water. Analyses were carried out between 150-700 nm to cover the whole spectrum. The optical density was mainly recorded at 600nm and at the λ_max of each dyestuff solution used following centrifugation of the withdrawn sample in a mini centrifuge for 5 minutes at 3,500 rpm. The percentage change in optical density D was then estimated for each dye soy broth solution as follows:

Where, O.D. is the optical density of reactive dye- soy broth solution before decolouoration with bacteria, O.D.

Dye Utilisation Studies

The decolouration of soy broth solutions containing dyes was investigated using Shewanella putrefaciens . Reactive Red and Reactive black solutions as well as real textile factory effluent was used. The performance of this bacteria to degrad reactive dyes was further tested against a number of other microorganisms known to biodegrad certain classes o azo compounds. The experiments were carried out using conical shake flasks containing a dye solution of known concentration, which were seeded by an inoculum of cells. Initial results (given in table 2) from these batch tests indicated that all dyes solutions were not biodegraded by these bacteria apart from Shewanella putrefaciens which showed exceptional colour removal abilities. It was also recognised that this culture may be responsible for the transformation of the dye molecule into a less conjugate form, thereby, facilitating colour removal.

Table 2 Percentage decolouration using various bacteria genera.

Visible/ UV Spectra

Figures 1-3 herewith show UV Visible scans for single component reactive dyes and textile effluent, from different textile factories at different discharge ports, before and after biodegradation using Shewanella putrefaciens . The peaks were mainly around 462, 566 and 600nm. The high absorbency in the UV spectrum of these effluents shows that the dyes also absorb light in this range, although some of this absorbency may be due to additional solvents and additives rather than the dyes themselves. After contact with the biosorption system for 24 hours the absorbency in the visible spectrum was reduced significantly due to the aerobic breakdown of the azo bond. Absorbency in the UV spectrum however is still quite high due to the UV absorbency found in the initial sample but also additional absorbency which could only have originated from the breakdown of the dyes. These results also show that the removal of colour is mainly by bacteria metabolism rather than colour bioadsorption on the cells as shown in the next section.

Adsorption Of Colour On The Biomass In some cases, utilisation of dye from the solution may become confused with physical adsorption onto the biomass thus reducing the bulk liquid concentration. Therefore, a series of experiments was undertaken to establish if the dyes were being degraded or simply adsorbed by the bacteria. This involved the sterilisation of bacteria cells followed by contacting the bacteria with dye effluent of a known concentration as described else where in this report. The results obtained indicated that colour removal by biosorption accounted for less than 1% of the total colour removed. Hence the effect of colour removal by biosorption onto the biomass can be neglected and the colour is said to be removed via degradation and breakage of the azo bond. Biological Activated Carbon (BAC)

Biological activated carbon is basically the combined effect of activated carbon adsorption and biological degradation of adsorbate species. In this work BAC systems were studied in batch and fixed bed systems. A series of experiments was carried out with batch reactor configuration in which textile effluent of a known dye concentration was contacted with the systems described in Table 3 below. The experiments were designed so as to compare the performance of different systems and to test the ability of the bacteria to grow on the COD available in the effluent without the addition of extra nutrients.

Table 3 experimental batch systems

The adsorption capacity of activated carbon and a support material was considered in systems 1 and 7 In general the results indicate that although colour was removed to a certain degree in all cases, cell immobilisation on activated carbon gives by far better performance than that with other support media or indeed free bacteria systems. The combination of bacteria and carbon, with the required amounts of essential nutrients added, resulted in colour reduction of around 95% while no significant change (22%) was noticed in the bacteria and support media systems. The colour reduction due to carbon adsorption was found to be around 77% in 24 hours and only 4% was removed by the supporting material which can be considered to have no adsorption ability towards reactive dye textile effluent. Furthermore, after an initial period of acclimatisation the GAC with immobilised cells reduced colour more effectively than the sterile carbon. The slightly lower rates of colour removal by the biological activated carbon system of these experiments is probably due to cells or nutrient broth residues blocking the macro-pores of the carbon, and the subsequent increase over conventional GAC a result of the combination of adsorption and biological degradation.

Carbon has a good adsorption capacity if contact time is sufficient enough, but can form a good support for the immobilisation of the bacteria and the combination gives an excellent colour removal compared to other support materials considered. When comparing the effect of colour reduction when variable amounts of nutrients are added, it can be seen that the bacterial activity in systems 6 and 10 resulted in 95% colour reduction.^* It is known that bacteria cells like all viable organisms must synthesise all chemicals needed to operate, maintain and reproduce the cells. This requires compounds for metabolism including fats and lipids, polysaccharides, and proteins which are provided in the nutrient broth in this case. The results reflected by these experiments shows that these compounds were not provided from the constituents of the textile effluent. In fact the dyestuffs act as a carbon and hydrogen source for the bacteria cells, therefore the additional compounds and elements needed for metabolism need to be added to the dye bacteria system. In this case the additional nutrients are added in the form of nutrient broth.

The Continuous Bio-Adsorption System The BAC columns operated were seeded by pumping the bacteria and culture in the liquid media, containing bacteria in their exponential growth phase. Turbidity measurements at the column inlet and outlet gave an indication of the amount of biomass being immobilised on the activated carbon and the support material. Initial results from these experiments are given in Figure 4 , in a plot of total number of bacteria adsorbed versus time. It was found that GAC appears to have the ability to act as a support for immobilised cells, as substantial number of bacteria are immobilised in these columns. The initial rate of immobilisation follows a decay curve due to bacterial saturation on the support material. The successful degradation of azo dyes by this bacterium is dependent upon the bacteria having adequate supplies of -nutrients . This was provided by the soy broth. To enhance the BAC column performance, a sterile nutrient broth was pumped through the bed before bacteria seeding to ensure adequate nutrient supply and column performance. Two systems were considered. The first system built was a continuous recycle, one column system (diameter 50mm & 100mm length) as illustrated in Figure 5 herewith. The system consists of two phases:

1. Aerobic ~ seeding the support packing with bacteria 2. Anaerobic ~ decolorisation of textile effluent

During the aerobic phase the Shewanella Putrefaciens was grown as described in the passage hereinbefore headed Culture Growth. The samples taken from the shake flasks were used to inoculate the 10 Litre aerobic reactor, which is filled with soya broth and temperature controlled at 30°C. The inoculated soya broth is then pumped upward through the glass column and on exit from the column is fed back to the aerobic reactor. Soya broth was supplied aseptically to the aerobic reactor at a pre-deter ined rate to insure maximum growth of the culture. When a sufficient biofilm has grown on the support material, the aerobic reactor was switched off and the second phase of the process began. The feed for the anaerobic phase was textile effluent from a local textile dyeing plant. The effluent was neutralised using hydrochloric acid and extra nutrients in the form of yeast extract was added. This mixture was then pumped through the glass column using a recycle system, the effluent was passed through the column several times. The results achieved from the system are displayed in terms of the difference of optical density in the column influent and effluent.

The results from this system showed that a once through, single column, as illustrated in Figure 5 had the potential to decolourise the textile effluent to an acceptable (limit of 0.6 at λmax) . Therefore the above described system was adapted so that the effluent was only passed through the column once. Results from the once through single column unit are displayed in Figure 6 herewith.

During process optimisation studies it was established that if the residence time of the textile effluent in the column was increased then there would also be an increase in breakthrough time (ie the life of the column), thus a second column (50mm diameter & 100mm length) was added in series (figure 8). The same seeding procedure as outlined above was still employed. The results are displayed in Figure 7 herewith.

The bioadsorption system maintained its colour removal ability for around three weeks.

Testing concentrated dye effluent

Experiments were extended to test the durability of the system and its ability to decolourise concentrated textile effluent. This will also result in reducing the size of the columns needed to treat the textile effluent on an industrial site. Trials were firstly completed using test-tubes, whereby several dilutions of the concentrated dye were placed in test-tubes, neutralised using hydrochloric acid, nutrients added and the mixture inoculated using bacteria. The test-tubes then stored in a shaker at 30°C for a given time period. The optical density of the tubes was measured at given time periods. From the test-tube results achieved, it has been shown that an acceptable rate of decolorisation was achieved using dilution factors as low as 1:10.

Alternative Nitrogen Sources Nutrient availability is essential for the growth and metabolism of the bacteria cells. The results reflected by these experiments show that these compounds were not provided from the constituents of the textile effluent. The results show that the additional nutrients are indeed required for a reasonable growth and hence colour reduction. Also the increase in nutrient concentration appears to be minimal due to sufficient concentration of compounds which the effluent alone cannot provide. This agrees well with the results obtained from studying the effect of broth concentration on the growth of this bacterium. In_. the shake flask batch systems inhibition was encountered after a period of 24 hours where little or no change in colour took place. However, the additional nutrient broth started the dye utilisation reaction once more. This may be attributed to the fact that the additional nutrients required for the metabolism and provided by the nutrient broth has been depleted, therefore halting any further growth. Also, with the shake flask experiments inhibition occurred due to the formation of detrimental by- products. In the biological activated carbon systems however, less retardation was evident (hence better colour removal) which may suggest that any detrimental by products formed may be adsorbed by the carbon therefore reducing the inhibitiye effect on the bacteria. Colour of blank, i.e., the reduction of colour due to the oxidation with no bacteria or carbon present was 11.5% in 24 hours. The colour where bacteria was seeded on effluent with no nutrients was reduced by 8% only and the bacteria growth activity resulted in a COD reduction of lOOmg/1. Replacement of Soya Broth

An investigation was carried out to determine an alternative growth medium for S. Putrefaciens. Soya broth which is currently being used to grow the microbe, is both expensive and high in COD. The investigation was begun by formulating a list of factors and medium components known to affect growth such as temperature, pH concentration of carbon source, minerals available etc. Then a series of investigations was conducted varying a single factor at a time to determine the optimum choice for each. The process is straight forward and is commonly used to discover the optimal growth medium for a given micro-organism or a particular process. Factors such as cost and, in this case, COD of the medium are important.

From the results achieved, it has been shown that approximately half the growth attained using 30g/L soya broth may be produced 94% cheaper (£1.17/L vs £0.066/L) by using the formula displayed in table 4 below.

Table 4 Growth media for Shewanella Putrefaciens

Toxicity

In the Daphnia immobilisation toxicity test procedure, groups of juvenile Daphnia magna (less than 24 hours old) are exposed to the sample diluted with reference freshwater, over a range of concentrations in an appropriate range will, under otherwise identical test conditions, exert toxic effects on the swimming activity (and survival) of Daphnia. These extend from an absence of effects at lower test concentrations to immobilisation of all daphnids at higher test concentrations.

Where appropriate, the EC₅₀, NOEC and LOEC were determined by statistical analysis using the ToxCalc® data processing package. The determination of the 24-h and 48-h EC₅₀ values is based on the proportion of immobilised Daphnia in the different test concentrations. Where the test response data did not permit determination of the EC50 value, the percentage immobility of daphnids was recorded.

Data obtained during the test were deemed to be valid; based on the absence of morality with the test controls, and measurement of the dissolved oxygen levels at the end of the test. The 48-h EC₅₀ value of the zinc reference toxicant was 1.40 mg Znl^-1 (based on analytically measured concentrations of zinc) . This value is within the limits defined for AQC of this procedure at this laboratory.

The 48-h Daphnia immobilisation toxicity test data for the pilot plant samples is shown in Table 5 below.

Sample Concentration Calculated 48- EC50 % NOEC LOEC

(% v/v sample) dilution of (% v/v sample) Daphnia ' pilot plant (95% confidence immobilisation effluent limits)

Red No. 1 32 1:4® 100 1:150 50.4 (40.3- 63.1) 100 32 100

Red No.2 32 1:469 0 100 1:150 >100 0 100 >100

Red No.3 32 1:469 0 100 1.150 >100 0 100 >100

Navy No. 1 32 1:469 0 100 1:150 56.6 (* - *) 100 32 100

Navy No.2 32- 1:469 0 100 1:150 >100 40 32 100

Navy No.3 32 1:469 100 1 150 60.3 (52.7-68.9) 90 32 100

Reference 1.30 mg Zn ¹ toxicant (1.08 - 1.56) The Table shows the measured toxicity in terms of the test concentrations prepared at WRc-NSF, and also in terms of the calculated dilution of pilot plant sample itself. The data for test concentrations 0, 1.0, 3.2 and 10% (v/v) sample (for which no effects were observed) have been omitted for the sake of clarity.

Only four of the samples tested caused immobilisation of the daphnids at the highest test concentration (Red Nol, Navy Nol, Navy No2 (partial response) and Navy No3) . No effects were observed at the next dilution of sample (32% v/v) .

The measured 48-h EC50 values were;

Red Nol = 50.4% Red No2 = >100% Red No3 = >100% Navy Nol = 56.6% Navy No2 = >100% Navy No3 = 60.3%

In this summary the following points are noted;

1. On this occasion the observed 48-h EC₅₀ values of the pilot plant samples lay between 32 and >100% v/v, that is between less than 1:150 dilution and 1:469 dilution of the effluent. 2. The minimum dilution required before toxicity exceeds 100% v/v has not yet been established for samples Red No2 and No3 and Navy No 2. 3. Consideration should be given to the choice of diluent in preparation of the pilot plant samples for any future toxicity testing. Filtered groundwater or de-chlorinated tap water would be a more appropriate choice of diluent to discriminate between apparent and intrinsic toxicity.

Claims

2. A process for the bioadsorption of a dye using a bioadsorbent material, wherein the bioadsorbent material is a dye-degrading bacterium on or in an adsorbent support material.

2. A process as claimed in Claim 1 wherein the bacterium is Bacillus Gordonae al . , Bacillus Beneovorans al . , Pseudomonas Putida or Pseudomonas sp.

3. A process as claimed in Claim 1 or Claim 2 wherein the bacterium is Shewanella Putrefacrens.

4. A process as claimed in any one of Claims 1-3 wherein the support medium is activated carbon.

5. A process as claimed in Claim 4 wherein the support medium is granulated activated carbon.

6. A process as claimed in Claim 4 or Claim 5 wherein the support material is one or more materials selected from the group comprising: F-400, ■ Centaur, EA207, C207, Chilean lignite carbon and lignite coke carbon.

7. A process as claimed in any one of the preceding Claims wherein the dye is an azo-dye.

8. A process as claimed in any one of the preceding Claims which involves treating the dye with a relatively short hydraulic retention time and relatively long solid retention time.

9. A process as claimed in any one of the preceding Claims which is a continuous fixed- bed bioadsorption process.

10. Apparatus for the bioadsorption of a dye comprising a passageway for the passage of the dye thereacross, . within which passageway is located a bioadsorbent material comprising a dye-degrading bacterium on or in an absorbent support material.

11. Apparatus as claimed in Claim 10 wherein the bioadsorbent material is as defined in any one of Claims 2 to 6.

12. Apparatus as claimed in Claim 10 or Claim 11 wherein the apparatus is a multistage arrangement.

13. Apparatus as claimed in Claim 12 wherein a first part of the apparatus is a continuous aerobic reactor, followed by a second part being a one or more columns in a fixed bed- configuration.

14. Apparatus as claimed in Claim 10 or Claim 11 wherein the apparatus is a cartridge assembly.

15. Apparatus as claimed in Claim 14 wherein the cartridge assembly is adapted to fit within a larger unit.

16. A method for the bioadsorption of a dye comprising passing the dye across a bioadsorbent material as defined in any one of Claims 2 to 6.

17. Use of a bioadsorbent material as defined in any one of Claims 2 to 6 in ths bioadsorption of a dye such as a reactive dye-

18. A process for the bioadsorption of a dye substantially as herein described and with reference to the drawings.