GB2439099A

GB2439099A - Method and apparatus for determining the biomass activity of a fluid

Info

Publication number: GB2439099A
Application number: GB0611589A
Authority: GB
Inventors: Freda Hawkes; Alan Guwy; Tim Patterson; Giuliano Claudio Premier
Original assignee: UGCS
Current assignee: UGCS
Priority date: 2006-06-12
Filing date: 2006-06-12
Publication date: 2007-12-19
Also published as: GB0611589D0

Abstract

A method of determining the biomass activity of a fluid carrying a biomass comprises (i) introducing the fluid carrying the biomass into a receptacle; (ii) removing fluid from the receptacle so as to increase the concentration in the receptacle of biomass from the fluid; (iii) introducing an oxidising agent into the receptacle so as to facilitate enzyme-catalysed release of oxygen; and (iv) measuring the oxygen produced in step (iii) to provide a measure of the biomass activity of the fluid. The concentration step (ii) may involve adding a flocculating or coagulating agent or subjecting the receptacle to a centrifuging action. Also shown is an apparatus for use in such a method.

Description

1 2439099 Method and apparatus for determining the biomass activity of

a fluid The present invention relates to a method and apparatus for determining the biomass activity of a fluid (especially, but not exclusively, aqueous fluids) The enzyme catalase is well-known for its ability to convert hydrogen peroxide to water and oxygen (see, for example, "Biochemistry", 5th Edition, Berg, Tymoczko and Stryer, published by W.H. Freeman and Co., New York, (2002) page 614) . Methods based on the reaction between catalase and the oxidant, hydrogen peroxide, have been developed to determine the organic content of a fluid. Those methods described in W096/18896 and US5863749 rely on the disruption of cells to release catalase, which may not give an accurate determination of the viable biomass of the fluid.

Furthermore, the methods described in W096/18896 and US5863749 do not readily permit low concentrations of organic content to be measured. The present invention addresses one or more of these problems.

In accordance with a first aspect of the present invention there is provided an apparatus for determining the biomass activity of a fluid carrying a biomass, the apparatus comprising a receptacle for performing a catalase-oxidant reaction, the receptacle being in user-operable fluid communication with a fluid inlet path for introducing the fluid into the receptacle, an oxidant inlet path for introducing an oxidant into the receptacle, and an oxygen outlet path, the oxygen outlet path being provided with a gas sensor for sensing oxygen produced by a catalase-oxidant reaction performed in the receptacle, the apparatus being provided with a means for increasing the concentration of the biomass in the receptacle.

The biomass may be associated with a solid, for example, the biomass may be attached to a solid. Alternatively, the biomass may be free-swimming i.e. planktonic biomass. The apparatus of the present invention may be suitable for determining the biomass activity of a fluid when the biomass is planktonic and/or when the biomass is associated with a solid.

The apparatus is preferably suitable for determining the biomass activity of a liquid carrying a biomass.

Biomass activity is equivalent to the degree of biomass respiration in a sample. This further reflects the amount of living biomass in a fluid. The biomass activity may typically be determined by determining the amount of catalase that is released or present in a sample; a large amount of catalase indicates high biomass activity. The amount of catalase present may be determined by adding an oxidant, such as hydrogen peroxide, to the sample. The catalase and hydrogen peroxide react to produce oxygen, the amount of oxygen being related to the amount of catalase present in the sample. The amount of oxygen produced is therefore indicative of the biomass activity of the sample.

The means for increasing the concentration of the biomass in the receptacle may be a means for increasing the concentration of a solid with which the biomass is associated.

The apparatus of the present invention facilitates the reaction between the oxidant and the catalase generated by, or present in, the biomass in the solid.

In use, the volume of the fluid is greater than the capacity of the receptacle. The means for increasing the concentration of the biomass in the receptacle facilitates the localization of biomass from fluids containing relatively low concentrations of biomass.

The oxidant inlet path may be provided with a source of oxidant. The fluid inlet path may be provided with a source of fluid.

The gas sensor may be a sensor that measures gas flow or gas concentration or gas pressure. The gas sensor may specifically detect the presence of oxygen, as opposed to any other gas. The gas sensor may separate oxygen from other gases, for example, by using the magnetic properties of oxygen.

Each of the fluid inlet path, oxidant inlet path and oxygen outlet path may be put in communication with the receptacle by, for example, providing user-operable valves. Each of said paths may be provided with a valve.

The means for increasing the concentration of biomass in the receptacle may comprise one or more of a filter, a means for adding flocculating or coagulating agent to the receptacle, a means for promoting evaporation of the fluid and a centrifuge with which the receptacle is associated.

It is preferred that the apparatus is further provided with a fluid outlet path for the egress of treated fluid from the receptacle. This allows large volumes of untreated fluid to enter the receptacle, permitting the collection of solid from the large volume of fluid. This is of particular use when the concentration of solid is low. The fluid outlet path may be isolatable from the receptacle. This is of particular use when oxygen is being generated in the receptacle so that oxygen does not flow through the fluid outlet path as opposed to the oxygen outlet path.

It is preferred that the apparatus is provided with a mixer for mixing the contents of the receptacle. The mixer may iS comprise a stirrer, such as a magnetic stirrer. The magnetic stirrer may comprise a rotatable stirring component for placement inside the receptacle and an associated stirring plate or housing for generating the magnetic field that causes the stirring component to rotate.

The apparatus may further comprise a pump for urging the oxidant to the receptacle, and may further comprise a pump for urging the fluid to the receptacle.

A common conduit may be provided for the introduction of fluid and oxidant into the receptacle. This may conveniently be provided by the fluid inlet path and oxidant inlet path meeting at a junction, the common conduit extending from the junction to the receptacle.

The apparatus may comprise two or more gas sensors. For example, this may facilitate the measurement of oxygen over a wider range of pressures than may be possible using only one sensor. Each of the two or more gas sensors may be a low pressure gas sensor such as a Honeywell 0-5 inch H20 pressure sensor(RS Components, Corby, UK), a high pressure gas sensor such as a DruckMPl4OO 0-ibar pressure transducer (RS Components, Corby, UK) or an oxygen concentration monitor such as a Sable Systems International FClb (Sable Systems International, Las Vegas, USA) It is preferred that one or more of the gas sensors is isolatable from the receptacle. It is further preferred that each of the gas sensors is individually isolatable from the receptacle. This would allow the user to selectively use the desired gas sensors, which may be of use if, for example, the pressure of oxygen generated was so low that it would not register on a high pressure sensor. Automatic selection of the desired gas sensors may also be achieved automatically, for example, under the instruction of a programmed control means, such as a programmed circuit board or computer.

In the event that the apparatus comprises a pump for urging the fluid to the receptacle, then the fluid inlet path may comprise a bypass conduit that allows fluid to be provided to the receptacle without use of said pump. This conduit may take the form of a bypass "loop" arranged so that one common fluid inlet may be used, irrespective of whether the pump is used.

The apparatus may comprise a fluid flow sensor. Such a fluid flow sensor may be provided in the fluid inlet path or a fluid outlet path, if provided.

In accordance with a second aspect of the present invention there is provided a method of determining the biomass activity of a fluid carrying a biomass, the method comprising (i) introducing the fluid carrying the biomass into a receptacle (ii) removing fluid from the receptacle so as to increase the concentration in the receptacle of biomass from the fluid (iii) introducing an oxidizing agent into the receptacle so as to facilitate enzyme-catalysed release of oxygen and (iv) measuring the oxygen produced in step (iii) to provide a measure of the biomass activity of the fluid.

The method of the present invention facilitates the measurement of the biomass activity of fluids carrying a relatively small concentration of biomass.

The biomass may be planktonic (i.e. free-swimming) in said fluid or the biomass may be associated with a solid which is carried by said fluid. The biomass may be attached to the solid.

The fluid is preferably a liquid, and is preferably an aqueous liquid.

In the method of the present invention, the total volume of fluid passing into the receptacle is greater than the capacity of the receptacle.

The fluid removed from the receptacle in step (ii) carries a lower concentration of biomass than the fluid introduced into the receptacle in step (i) . It is preferred that the fluid removed from the receptacle is substantially devoid of biomass. If the biomass is associated with a solid, it is preferred that the fluid removed from the receptacle in step (ii) carries a lower concentration of solid than the fluid introduced into the receptacle in step (i) . It is further preferred that the fluid removed from the receptacle is substantially devoid of solid.

The method may further comprise isolating biomass from the fluid prior to step (ii) . This causes the biomass to separate locally from the fluid, effectively extracting the biomass from the fluid, thus decreasing the concentration of the biomass carried by the fluid, allowing the fluid (carrying its lower concentration of biomass) to be removed from the receptacle. If the biomass is associated with a solid, the method may comprise isolating solid from the fluid prior to step (ii) . This causes the solid to separate locally from the fluid, effectively extracting the solid from the fluid, thus decreasing the concentration of the solid carried by the fluid, allowing the fluid (carrying its lower concentration of solid) to be removed from the receptacle.

This isolation may be achieved by jntroduciflg a focculating or coagulating agent into the receptacle. 1ternatively or additionally, this isolation may comprise subjecting the contents of the receptacle to a centrifuging action.

The step of measuring the oxygen produced may comprise measuring the volume of oxygen produced. Alternatively or additionally, the step of measuring the oxygen produced may comprise measuring the pressure of oxygen produced. This may conveniently be achieved using a gas pressure sensor.

Furthermore, alternatively or additionally, the flow rate of gas may be measured (for example, using a flow meter) Alternatively or additionally, the step of measuring the oxygen may comprise measuring the time taken for a pre- determined volume of gas to be collected or for a pre-determined pressure of gas to be achieved. This may involve collecting gas in a space of given volume, and measuring the time taken for the gas pressure in the given volume to reach a pre-determined level. This latter step may be repeated at a frequency dependant on the volume of gas produced.

Step (ii) may comprise passing the fluid through a filter or evaporating fluid. Filtration in particular provides an effective way of concentrating biomass (and particularly biomass-carrying solid) in the receptacle.

The receptacle may be provided with a fluid inlet for the ingress of solid-bearing fluid and a fluid outlet for the egress of treated fluid. This allows large volumes of fluids to be treated; this is particularly useful if the biomass activity of a fluid is low.

The contents of the receptacle may be stirred prior to the addition of oxidizing agent into the receptacle and/or during enzyme-catalysed release of oxygen. This helps to promote the reaction between catalase released by the biomass and the oxidant.

The method may comprise providing an apparatus for determining the biomass activity of a fluid, the apparatus comprising the receptacle, a fluid inlet path for introducing the fluid into the receptacle, an oxidant inlet path for introducing the oxidizing agent into the receptacle and an oxygen measurement path for permitting the measurement of oxygen.

The fluid inlet path, the oxidant inlet path and the oxygen measurement path may be isolatable from one another and the receptacle. The apparatus may be provided with an outlet path for the egress of treated fluid from the receptacle.

The outlet path may be isolatable from the fluid inlet path, the oxidant inlet path, the oxygen measurement path and the receptacle.

Prior to step (ii), the oxidant inlet path and oxygen measurement path may be isolated from the receptacle.

In this case, after step (iii) and prior to step (iv), the fluid inlet path (and optionally the outlet path) may be isolated from the receptacle, the oxidant inlet path being de-isolated from the receptacle.

After step (iii) and prior to step (v) (and preferably prior to step (iv)), the oxygen measurement path may be de-isolated from the receptacle.

The oxidant may comprise hydrogen peroxide.

Steps (i) and ii) may be performed in parallel i.e. simultaneously. One or both of step (i) and step (ii) may be repeated. One or both of steps (i) and (ii) may be continuous. Alternatively, fluid Carrying solid may be introduced into the receptacle (and/or fluid removed from the receptacle) in one or more batches.

The method of the second aspect of the present invention may comprise the use of the apparatus of the first aspect of the present invention.

The present invention will now be described by way of example only with reference to the accompanying drawing: Figure 1 is a schematic representation of an embodiment of an apparatus in accordance with the first aspect of the present invention.

Figure 1 shows a schematic representation of an embodiment of an apparatus for determining the biomass activity of a fluid suspected of carrying biomass in accordance with the first aspect of the present invention. The apparatus is shown generally by reference numeral 1 and comprises a receptacle 28 in user-operable fluid communication with a fluid inlet path 2 for introducing the fluid into the receptacle, an oxidant inlet path 7 for introducing an oxidant into the receptacle and an oxygen outlet path 12, the oxygen outlet path being provided with a gas sensor 16, 17, 20, the apparatus being provided with a means for increasing the concentration of biomass in the receptacle, the means being provided in the form of a filter 31. In this case, the receptacle 28 is a filter housing (Classic Filters PP2211.461 filter housing, Classic Filters Ltd., Rochester, Kent, UK) and the filter 31 is a filter for liquids, the filter having a pore size of 0.3 microns made of bonded borosilicate glass microfibres (Classic Filters 25.64.2E filter, Classic Filters Ltd., Rochester, Kent, UK) A pump 4 for urging fluid (not shown) from fluid inlet 3 to the receptacle is provided in the fluid inlet path 2. In the present case, the pump is a 0.9 horse power centrifugal pump. A bypass conduit 5 is provided so that fluid may be transferred to the receptacle 28 from the fluid inlet 3 without use of the pump 4. This may be desired, for example, if the pressure or velocity of fluid is sufficient to enable transfer of fluid to the receptacle without the use of the pump 4. A user-operable valve 6 is provided to isolate the fluid inlet 3 and pump 4 from the receptacle 28.

A pump 9 for urging oxidant (not shown) from a source of oxidant 8 to the receptacle 28 is provided in the oxidant inlet path 7. In this case, the pump is a micropump. A user-operable valve 10 is provided to isolate the source of oxidant 8 and pump 9 from the receptacle 28.

The fluid inlet path 2 and oxidant inlet path 7 meet at a junction from which extends common conduit 11 to the receptacle 28. This allows both the fluid and oxidant to flow (generally one at a time) into one receptacle inlet 32.

Receptacle 28 has a generally cylindrical shape, and has a receptacle inlet 32 and outlet 33 located near the top. The receptacle inlet 32 is in user-operable fluid communication with the fluid inlet 3 and the source of oxidant 8 so as to be able to receive the fluid and oxidant as required. The receptacle inlet 32 also acts as an outlet for oxygen, and is in user-operable communication with the oxygen outlet path 12. The receptacle outlet 33 is in user-operable communication with fluid outlet path 24 and therefore fluid outlet 27.

The receptacle 28 is provided with a filter 31 for concentrating any biomass-carrying solid (not shown) carried by the fluid. The filter 31 is arranged in relation to the receptacle outlet 33 so that fluid may not pass from the receptacle to the receptacle outlet 33 without passing through filter 31. A rotatable stirring component 29 is provided in the receptacle 28. The stirring plates 30 cause the stirring component 29 to undergo a rapid rotational motion, thus stirring the contents of the receptacle 28.

A sensor arrangement (not shown) is used to determine whether the filter 31 is in place in the receptacle 28. The sensor arrangement is in communication with the pump 4 so that the pump 4 is inoperable in the event that the sensor arrangement determines that the filter 31 is not in place in the receptacle 28.

Fluid outlet path 24 is provided with a flow sensor 26 to measure the flow of treated fluid from the receptacle outlet 33. In this case, the flow sensor is a turbine flowmeter (Cole Palmer, Vernon Hills, Illinois, USA) . Fluid outlet path 24 is also provided with a water flow detector (not shown) . The water flow detector detects the presence of water; when the apparatus is first operated, air may be pumped through the fluid outlet path 24, causing the flow sensor 26 to sense a flow. The water flow detector will give an indication as to when the flow through the fluid outlet path is caused by water, not air.

Oxygen outlet path 12 is provided with three gas sensors in the form of high pressure gas sensor 17, low pressure gas sensor 20 and oxygen concentration monitor 16. Valves 15, 18 are provided to allow the isolation of the oxygen concentration monitor and the low pressure gas sensor 18 respectively from the receptacle 38. A further valve 13 is provided to isolate all three of the low pressure gas sensor 20, high pressure gas sensor 17 and oxygen concentration monitor 16 en bloc from the receptacle. An assay chamber 19 is provided in the flow path between the valve 18 and low pressure gas sensor 20 to accumulate oxygen. A valve 21 is provided downstream of the low pressure gas sensor 20 to control egress of gas through exhaust 23. A carbon filter may be fitted to the exhaust to treat exhausted gases if required. A particle filter (not shown) is placed immediately downstream of the high pressure gas sensor 17 to prevent damage to the sensor that may be caused by particulate matter. Further particle filters (not shown) are placed in the flow path between valve 18 and the assay chamber 19 and immediately upstream of the low pressure gas sensor 20. An expansion chamber 14 is provided immediately downstream of the valve 13 to provide an expansion space, should the oxygen-generating reaction in the receptacle 28 generate a large volume of gas. In this case, the assay chamber 19 and expansion chamber 14 are provided by filter housings (Classic Filters Ltd., Rochester, Kent, UK) The method of operation of the apparatus 1 will now be described with reference to Figure 1. Valves 10, 13 are closed and valves 6, 25 opened (if the valves are not in the correct respective positions) . This allows fluid suspected of containing biomass to be delivered to receptacle 28 and treated fluid to be removed from the receptacle 28 without any fluid being delivered to the oxidant inlet path 7 or the oxygen outlet path 12. Fluid inlet 3 is placed in fluid communication with a fluid source (not shown) and pump 4 urges fluid to the receptacle 28 via receptacle inlet 32.

Fluid passes to receptacle outlet 33 via filter 31, the filter 31 being arranged to ensure that fluid cannot pass to the receptacle outlet 33 without passing through the filter 31. Particles larger than the filter pore size will not pass through the filter 31 and will be collected in receptacle 28. Filtered fluid passes out of the receptacle outlet 33 via valve 25 to the fluid outlet 27. The flow rate of filtered fluid is measured using flow sensor 26. The flow sensor may be in communication with the pump 4 (for example, via a microprocessor or the like) so that if the flow rate is lower than or equal to a pre-determined level then the pump 4 is turned-off or, alternatively, the output of pump 4 increased so that the flow rate is increased.

Once the desired sample size has been obtained in the receptacle 28 (or fluid has been passed through the receptacle 28 for a suitable time) then pump 4 is turned-off and valves 6, 25 are closed. It may be desirable to leave-open valve 25 for a short period after closing valve 6 to allow excess fluid to drain under the influence of gravity from receptacle 28.

Valves 10 and 13 are opened, and pump 9 urges oxidant from the source 8 via the common conduit 11 to the receptacle 28.

Valve 10 is then closed. The magnetic stirrer is then activated to stir the contents of the receptacle 28. This brings the biomass (if present) effectively into contact with the oxidant, thus facilitating the release of oxygen resulting from the catalase-oxidant reaction.

Oxygen released from the reaction passes into expansion chamber 14. In the event that the user wishes to use the high pressure gas sensor 17 then valves 15, 18 are closed, thus directing substantially all of the oxygen to the high pressure gas sensor. After a pre-determined time (or if the pressure of the gas rises beyond a pre-determined level) valves 18, 21 are opened, allowing gas to vent to the atmosphere via exhaust 23.

In the event that the user wishes to use the low pressure gas sensor 20 then valve 18 is opened and valve 21 is closed. Gas passes into the assay chamber 19, and the gas pressure is measured using the low pressure gas sensor 20.

When the pressure reaches a pre-determined value as determined by the low pressure gas sensor 20, valve 18 is closed and valve 21 opened, allowing the contents of the assay chamber 19 and associated conduits (not shown) to vent to atmosphere via exhaust 23. The time taken for the gas pressure to reach the pre-determined value is recorded and represents a measure of the rate of production of a given volume of gas (a longer time indicating a slower rate of production of gas) . The rate of production of oxygen is dependent on, inter alia, the concentration of catalase; the rate of production of oxygen is generally proportional to the concentration of catalase. The concentration of catalase is dependent upon the biomass activity of the solid, and so the rate of production of oxygen is indicative of the biomass activity of the solid.

Valve 21 is then closed, valve 18 is opened, and the time taken for the gas pressure to reach its pre-determined value is once again recorded.

The measurement-venting process is repeated as often as desired dependant upon the volume of gas produced.

The apparatus 1 is arranged so that the high pressure gas sensor 17 may be used to monitor gas pressure whilst the low pressure gas sensor 20 is being used. In the event that the gas pressure measured by the high pressure gas sensor 17 is above a pre-determined level, then the apparatus would automatically open valves 18, 21 to allow gas to vent to atmosphere.

Measurements may be taken using the oxygen concentration monitor 16 by operation of valve 15.Such a monitor would specifically monitor oxygen concentration, which may be advantageous if it is suspected that gases other than oxygen are being produced. Gases other than oxygen would be measured by a gas pressure sensor (such as high pressure gas sensor 17) and may therefore give misleading readings of oxygen evolved from the receptacle.

Once the desired measurements have been taken on the gas released from the receptacle 28, then valves 13, 18 and 21 are opened to allow the venting-off of any further pressure increase. Valve 25 is opened to drain fluid from the receptacle.

The filter 31 may be removed and cleaned, if desired. The receptacle 28 may be removed and emptied, if desired.

Example

A simple experiment was conducted to further demonstrate the method of the present invention. A soil sample suspected of containing biomass was air dried, crushed in a pestle and mortar and passed through an 85 mesh sieve. Material passing through the sieve was collected and weighed (l3.75g) These soil fines were suspended in approximately 16 litres of water in a header tank, giving an approximate suspended solids concentration of about O.85g/l.

An air-dried Classic Filters 25.64.3E filter was placed inside a Classic Filters PP211.46l filter housing. The header tank was then connected to the inlet of the filter housing and a pump used to pump the suspension from the header tank to the filter housing. Treated water (having passed through the filter) passed through the outlet of the filter housing. The filter housing was further provided with an outlet pipe for the outlet of oxygen, the outlet pipe extending into a vessel containing water.

Once sufficient sample had been collected, the pump was turned-off and the filter housing isolated from the pump. A magnetic stirring arrangement similar to that described above was used to mix the contents of the filter housing. An oxidant (lOml of 3% hydrogen peroxide) was added to the contents of the filter housing by syringe. Addition of the oxidant caused the evolution of a slow but steady flow of bubbles from the end of the outlet pipe (about 1 bubble every 1 to 2 seconds), indicating that a catalase-oxidant reaction was occurring between the biomass in the filter housing and the oxidant. Bubbles were evolved over a period of about 30 minutes.

Once the evolution of bubbles had been completed the solids remaining in the filter housing were extracted and weighed.

Approximately 4.5g of solid had been recovered from the 6 litres of fluid that had been pumped through the filter in a minute period.

The system was initially operating at a pressure of about 1 bar, although this decreased to 0.6 bar at the end of the test. The pumping rate through the system was also found to decrease through the test, possibly due to a reduction in -pressure and/or fouling of the filter.

This simple experiment illustrates that it is possible to isolate and collect solid sample from a fluid sample in a receptacle and use that same receptacle to perform an enzyme-catalysed oxygen-generating reaction.

Whilst the present invention has been described and illustrated with reference to a particular embodiment, it will be appreciated by those of ordinary skill in the art that the invention lends itself to many different variations not specifically illustrated herein. For that reason, reference should be made to the claims for determining the true scope of the present invention.

The apparatus of the present invention need not, of course, be manually operated; it could be automatic or semi-automatic. The apparatus could be provided with a. control unit or microprocessor for controlling the apparatus. The control unit may, for example, control the operation of the pump for urging fluid to the receptacle and/or the pump for urging oxidant to the receptacle. The control unit may control one or more of the valves, and may further control the operation of the stirring plates. Data generated by the apparatus could also be automatically collected andanalysed using a microprocessor. These data could be used to affect a change in the process which the apparatus is monitoring.

Claims

Claims 1. A method of determining the biomass activity of a fluid

carrying a biomass, the method comprising (i) introducing the fluid carrying the biomass into a receptacle (ii) removing fluid from the receptacle so as to increase the concentration in the receptacle of biomass from the fluid (iii) introducing an oxidizing agent into the receptacle so as to facilitate enzyme-catalysed release of oxygen and (iv) measuring the oxygen produced in step (iii) to provide a measure of the biomass activity of the fluid.

2. A method according to claim 1 wherein step (ii) comprises passing the fluid through a filter or evaporating fluid.

3. A method according to claim 1 or claim 2 wherein the fluid removed from the receptacle in step (ii) is substantially devoid of biomass.

4. A method according to any preceding claim further comprising isolating biomass from the fluid prior to step (ii) 5. A method according to claim 4 wherein isolating the biomass from the fluid comprises adding a flocculating or coagulating agent into the receptacle or subjecting the contents of the receptacle to a centrifuging action.

6. A method according to any preceding claim wherein the step of measuring the oxygen produced comprises measuring the volume of oxygen produced.

7. A method according to any preceding claim wherein the step of measuring the oxygen produced comprises measuring the flow rate of oxygen produced and/or measuring the oxygen produced comprises measuring the concentration of oxygen produced.

8. A method according to any preceding claim wherein the step of measuring the oxygen produced comprises measuring the time taken for a pre-determined volume of oxygen to be produced.

9. A method according to any preceding claim wherein the step of measuring the oxygen produced comprises measuring collecting oxygen in a space of a given volume and measuring the time taken for the gas pressure in the given volume to reach a pre-determined level.

10. A method according to any preceding claim wherein steps (i) and (ii) are performed in parallel.

11. A method according to any preceding claim wherein one or both of steps (i) and (ii) are repeated.

12. A method according to claim 11 wherein fluid carrying biomass is introduced into the receptacle (and/or fluid removed from the receptacle) in one or more batches.

13. A method according to any preceding claim wherein one or both of steps (i) and (ii) are continuous.

14. A method according to any preceding claim wherein the receptacle is provided with a fluid inlet for the ingress of fluid carrying biomass and a fluid outlet for the egress of treated fluid.

15. A method according to any preceding claim wherein the contents of the receptacle are stirred prior to the addition of oxidizing agent into the receptacle and/or during enzyme-catalysed release of oxygen.

16. A method according to any preceding claim comprising providing an apparatus for determining the biomass activity of a fluid, the apparatus comprising the receptacle, a fluid inlet path for introducing the fluid into the receptacle, an oxidant inlet path for introducing the oxidizing agent into the receptacle and an oxygen measurement path for permitting the measurement of oxygen.

17. A method according to claim 16 wherein the fluid inlet path, the oxidant inlet path and the oxygen measurement path are isolatable from one another and the receptacle.

18. A method according to claims 16 and 17, the apparatus being provided with an outlet path for the egress of treated fluid from the receptacle.

19. A method according to claim 18 wherein the outlet path is isolatable from the fluid inlet path, the oxidant inlet path, the oxygen measurement path and the receptacle.

20. A method according to claims 17 to 19 wherein, prior to step (ii), the oxidant inlet path and oxygen measurement path are isolated from the receptacle.

21. A method according to claim 20 wherein, after step (iii) and prior to step (iv), the fluid inlet path (and optionally the outlet path) is isolated from the receptacle, the oxidant inlet path being de-isolated from the receptacle.

22. A method according to claims 20 and 21 wherein, after step (iii) and prior to step (v), the oxygen measurement path is de-isolated from the receptacle.

23. A method according to claims 22 wherein, after step (iii) and prior to step (iv), the oxygen measurement path is de-isolated from the receptacle.

24. An apparatus for determining the biomass activity of a fluid carrying a biomass, the apparatus comprising a receptacle for performing a catalase-oxidant reaction, the receptacle being in user-operable fluid communication with a fluid inlet path for introducing the fluid into the receptacle, an oxidant inlet path for introducing an oxidant into the receptacle and an oxygen outlet path, the oxygen outlet path being provided with a gas sensor for sensing oxygen produced by a catalase-oxidant reaction performed in the receptacle, the apparatus being provided with a means for increasing the concentration of biomass in the receptacle.

25. An apparatus according to claim 24 wherein the means for increasing the concentration of biomass in the receptacle comprises one or more of a filter, a means for adding flocculating or coagulating agent to the receptacle, a means for promoting evaporation of the fluid or a centrifuge with which the receptacle is associated.

26. An apparatus according to claim 24 or claim 25 further provided with a fluid outlet path for the egress of treated fluid from the receptacle.

27. An apparatus according to any one of claims 24 to 26 provided with a mixer for mixing the contents of the receptacle.

28. An apparatus according to claim 27 wherein the mixer comprises a stirrer.

29. An apparatus according to claim 28 wherein the mixer comprises a magnetic stirrer.

30. An apparatus according to any one of claims 24 to 29 further comprising a pump for urging the oxidant to the receptacle and a pump for urging the fluid to the receptacle.

31. An apparatus according to any one of claims 24 to 30 comprising a common conduit for introduction of fluid and oxidant into receptacle.

32. An apparatus according to any one of claims 24 to 31 comprising one or more gas sensors.

33. An apparatus according to claim 32 wherein each of the two or more gas sensors is a low pressure gas sensor, a high pressure gas sensor or an oxygen concentration monitor.

34. An apparatus according to claim 32 or 33 wherein one or more of the gas sensors is isolatable from the receptacle.

35. An apparatus according to any one of claims 24 to 34 comprising a pump for urging the fluid to the receptacle, wherein the fluid inlet path comprises a bypass conduit that allows fluid to be provided to the receptacle without use of said pump.

36. An apparatus according to any one of claims 24 to 35 further comprising a fluid flow sensor.

37. A method of determining the biomass activity of a fluid substantially as hereinbefore described with reference to Figure 1.

38. An apparatus for determining the biomass activity of a fluid substantially as hereinbefore described with reference to Figure 1.