DE4332163A1 - Method and device for pollutant analysis in water samples - Google Patents

Abstract

A water sample to be analysed is mixed in a mixing vessel (5) with an algae suspension (8) withdrawn from a pure storage vessel (1). After a predetermined reaction time, the mixture is transferred into a measuring cuvette (29) and exposed to weak and modulated measurement light as well as to stronger and unmodulated photosynthetically active continuous light. As a function of the continuous light, the behaviour of the oxygen development of the fluorescence signals emitted by the mixture is detected. The measured values of the fluorescence and of the oxygen development are compared with each other in a computer (33) as a basis for the pollutant analysis. <IMAGE>

Description

The invention relates to a method and an apparatus for pollutant analysis of water samples with algae as an indicator goal means.

Algae have proven to be cost effective indicators for the Proven pollutant analysis of water samples. In addition, by suitable temperature control and nutrient or CO₂ addition to the Algae suspension any life or shelf life of the indicator approach can be achieved. In the well-known process for the analysis of pollutants using algae as an indicator The water samples spiked with algae suspension previously irradiated with continuous light. The intensity of the Ge mixed emitted fluorescent light represents a measure of that Pollution of the water samples. The disadvantage of this The method consists in that stray light and both fluorescent substances in water as well as from dead Chlorophyll-containing organisms emit light with the Fluorescent light is detected. An exact determination of the "Pure fluorescence yield" of the indicator organisms is not or only possible with difficulty. The procedure is therefore relatively un accurate and prone to interference from outside light.

The invention provides a remedy here. You have the task based on the determination of the pollution of water bodies to make it more reliable with limited effort.

In terms of the method, the solution to this problem according to the invention is
that an algae suspension is formed;
that a water sample is mixed with the algae suspension and is brought into contact with the algae sample over a predetermined reaction time;
that the mixture is irradiated with a weak and modulated first light signal (measurement light) and with a photosynthetically active, stronger but unmodulated second light signal (continuous light);
that, depending on the continuous light, the course of the oxygen development and the fluorescence signals emitted by the mixture is detected; and
that the measured values of fluorescence and oxygen development are compared as the basis of the pollutant analysis.

The device for pollutant analysis of water samples with Al According to the invention, gene as an indicator agent is characterized by this from that a water sample line and an algae suspension supply line can be coupled to a mixing vessel; that a pro benentnahmevvorrichtung for taking mixture samples from the Mixing vessel can be connected to a measuring cell; and that the Measuring cuvette at least one light source, an oxygen detector onsmittel and a light detector are assigned.

By capturing two of the photosynthesis relevant parameters - the fluorescence and the oxygen development lung - it is possible to check the physiological state of the mixture to grasp more precisely. Since the physiological process of Photo Synthesis of algae very sensitive to water Reacts pollutants, thus becomes an accurate pollutant analysis enables.

The measuring method according to the invention is based on the separation of fluorescence measurement and photosynthesis excitation light. First this Separation enables the combined measurement of fluorescence and oxygen signals. The combined capture of this photo The synthesis parameter increases the reliability of the measurement results nits.

The use of modulated fluorescence measurement light increased the measuring accuracy, since only the fluorescence yield is measured by photosynthetically active organisms. The white The modulated fluores received by the measuring amplifier can test signal of the test algae without the influence of incident stray light and interference fluorescence can be detected.  

A major advantage of the measuring system is the fact that complex signal patterns are recorded and that these Character pattern characterized by different pollutants be changed in different ways. This can be different from usual biological effects tests in addition to the simple regi effect of an effect also a reference to the few pollutants are given that cause the effect Has.

In a further development of the invention, not only the physiological process of photosynthesis, but also the dis Similation (breathing) of the algae is monitored. This is the course oxygen evolution in the absence of photosynthetic active light detected. Different types of pollutants inhibit this difference between two different physiological processes strong. For some pollutants, primarily the Electron transport in the cells disrupted, d. H. the photosyn these inhibited. Other toxides mainly damage the mitro chondria and can do better with inhibited dissimilation be recognized. Combined monitoring of dissimilation and assimilation can increase accuracy and analysis further improve the type of pollutant.

A short measuring time of only 6 Mi is also advantageous grooves. In a special embodiment of the measuring cell, at which is the measurement of fluorescence intensity and oxygen Development in two spatially separated measuring chambers at the same time tig, a measurement time of only 4 is sufficient Minutes. With a single measuring arrangement you can in a short time intervals of water samples for pollutant loads be searched. The upstream exposure time of Pollutants on the algae species added to the water sample sion is only about 30 minutes. So it succeeds in pollutant Analyze quasi continuously (every 6 or 4 minutes) and at only a small time delay with a single measurement to carry out order.  

The device for pollutant analysis according to the invention is special stable and robust and therefore for mobile use also suitable on unpaved waterways.

Automation is a further development of the invention of the measuring method in that measured values are periodic recorded, recorded by a computer, stored and sent to a Headquarters are transferred. Anything can be done in this central many measuring points monitored by a single operator become. So the pollutant analysis of water can not only more precisely, but also faster and more economically leads.

Advantageous further developments and refinements of the Erfin are marked in the subclaims.

In the following the invention based on two in the drawing tion schematically illustrated embodiments he purifies. The drawing shows:

Figure 1 shows the basic structure of a device for pollutant analysis according to the invention in a first embodiment.

Fig. 2 is a schematic representation of a measuring cell with two separate measuring chambers of an inventive device for pollutant analysis in a second imple mentation form

Fig. 3 is a diagram showing the temporal course of the fluorescence intensity and the development of oxygen during a measuring cycle depending on the speed of various light signals acting on the mixture; and

Fig. 4 is a diagram showing the course of the basic fluorescence intensity with regularly repeated measurements on Rhine water and change in the basic fluorescence when polluted.

Fig. 1 shows the essential components of a first exemplary embodiment from the pollutant analyzer. A temperature-controlled algae suspension storage vessel 1 is provided with a device for determining the optical density (refractometer) and a CO₂ feed line. A turntable 3 serves as a carrier for several mixing vessels 5 . A predetermined amount of a mixture of algae suspension 9 and a water sample is fed to each mixing vessel 5 with the aid of a conveyor device 7 designed as a pump block. The aliquot of the Algensus pension 9 is supplied via a suction line 11 from the Algenspensi ons storage vessel 1 and test water to the conveyor 7 via a line 13 from a valve 15 . In the exemplary embodiment shown, the valve 15 has two inlets 17 and 19 which, depending on the valve position, are connected to the valve outlet opening into the line 13 . In each of these two positions, the other inlet is closed.

Each mixing vessel 5 is a z. B. ausgebil Dete stirring device 21 assigned as a magnetic stirrer, by means of which the mixture introduced from the conveyor 13 into the mixing vessel 5 is intimately mixed. In a position A the mixing vessel is filled. A mixture sample for the actual pollutant analysis measurement can be taken in a position B of the mixing vessel that is 180 ° offset from position A. A height-adjustable suction line 23 is arranged at point B and can be immersed in the mixing vessel. When the valve 25 is open, the mixture sample can be sucked out of the mixing vessel by a peristaltic pump 27 and passed into a measuring cell 29 . The measuring cell 29 is provided with an overflow 31 . The measuring cell is assigned at least one light source, an oxygen detection means and a light detector. When light signals are irradiated into the measuring cuvette filled with the mixture sample, the fluorescence intensity is measured on the one hand by the light detector and on the other hand the oxygen development is measured by the oxygen detection means.

On the line between the measuring cell 29 and the Ven til 25 , a flushing device is switched on. To the Spülvor direction include a water reservoir 35 , two valves 37 , 39 and a wastewater vessel 41st The connection between Vorratsge vessel 35 and measuring cell 29 is made by opening the valve 37 ago. The rinsing water is discharged from the measuring cell 29 via the valve 39 into the waste water container 41 .

The basic process flow of the pollutant analysis is explained below using the exemplary embodiment according to FIG. 1. The process sequence requires at least two measuring cycles, of which a first (reference measuring cycle) serves to calibrate the analytical device using practically pollutant-free reference water (line 19 ) and a subsequent measuring cycle (analytical measuring cycle) with a polluted water sample supplied via line 17 to valve 15 is carried out.

During the reference measurement cycle, the algae suspension is fed via the suction line 11 and reference water from the valve inlet 19 via the valve outlet and the line 13 to the mixing vessel 5 . It was found that a mixing ratio of 6: 2 between reference or sample water and algae suspension is particularly suitable for the desired pollutant analysis. An algae suspension reference mixture amount of 8 ml ensures sufficient measuring accuracy for the pollutant analysis. After filling the mixing vessel at point A, the reference water is allowed to act on the algae suspension over a reaction time of, for example, 30 minutes. During this reaction time, the mixing vessel 5 is transported on a circular path through 180 ° to point B. There, the mixture is pumped into the measuring cell 29 via the height-adjustable, flexible line 23 with the valve 25 open using the hose pump 27 . The valves 37 and 39 are closed. Since the volume of the measuring cuvette, in this case for example 6-7 ml, falls below the volume of the mixing vessels (8 ml), the measuring cuvette is completely and additionally filled with part of the overflow 31 . Then, depending on the light signals radiated onto the sample, the fluorescence intensity and the oxygen development are measured over a period of, for example, 3 minutes and evaluated in a data processing device 33 .

After the measuring cycle the valve 25 is closed and valve 39 opened so that the contents of the cuvette 29 can be pumped via the hose pump 27 into the waste water vessel 41st Thereafter, the valve 39 is closed and the two valves 25 and 37 are opened. In this valve position, the measuring cuvette 29 and the mixing vessel at point B are filled with water from the water storage vessel 35 for rinsing. After the valve 37 has been closed and the valve 39 has been opened again , the rinsing water from the measuring cell 29 and the mixing vessel is also pumped into the waste water vessel 41 at point B.

The next mixing vessel is then transported to position B and a new mixture sample is taken for subsequent measurement in the measuring cell 29 , etc.

The reference cycle is used to increase the measuring accuracy for example five times with unpolluted reference water again get. The individual measured variables are averaged and as with ted reference pattern saved. Signal pattern from eating pollutant analyzes are at least with this one Reference patterns compared.

In order to be able to make a more precise statement about the amounts of pollutants and the type of load, measurements are carried out on loaded reference water after the reference measurements on unloaded reference water. The measured quantities which were measured during the measurements on the reference water samples loaded with different pollutants and different amounts of pollutants are also stored by the data processing device 33 and used as further reference patterns for comparison with signal patterns from subsequent pollutant analyzes of water samples. After a number of reference cycles with different loaded reference samples adapted to the respective measuring circumstances (ie to the expected types of pollutants) and to the desired measuring accuracy, the water sample analysis begins.

The subsequent measurement procedures under analysis conditions correspond to those of the previously described measurement under reference conditions, with the only exception that the valve 15 is switched to the water sample inlet 17 . Instead of the reference water to be examined, the contaminated Ge water sample is introduced into a mixing vessel 5 , which is located at point A. In a reaction time of approx. 30 minutes, the mixing vessel is transported to point B, where - as described above - the measuring cell is filled and the fluorescence intensity and oxygen measurement is carried out. The measured signals are transmitted to the data processing device 33 and evaluated there by comparison with the stored reference patterns.

FIG. 2 shows a schematic illustration of a measuring cell in an embodiment in which the measuring space is divided into two measuring chambers 51 and 52 connected via a capillary 53 . The measuring cell 29 is filled with the aid of the feed line 28 via the lower measuring chamber 52 . In this embodiment, only two mixing vessels 5 are provided, which are firmly attached and can alternatively be filled via valves or emptied to fill the measuring cell. In this embodiment too, an overflow 31 is provided above the upper measuring chamber 51 .

The development of oxygen as a function of photosynthetic light radiated onto the sample is measured in the lower measuring chamber. For this purpose, light is irradiated 55 perpendicular to the right Küvettenachse to the sample mixture in the lower measuring chamber via an optical waveguide. An oxygen electrode 57 is immersed in the measuring chamber on the opposite side and it detects the photosynthetically released amount of oxygen. Via a capillary 53 , the lower measuring chamber 52 is connected to the upper measuring chamber 51 so that a liquid exchange between the two measuring chambers is possible, but the light signals in the two chambers are decoupled. In the upper measuring chamber 51 , the fluorescence of the mixture is measured as a function of different light signals radiated onto the sample. For this purpose, weak, modulated measuring light is emitted onto the contents of the upper measuring chamber via an emitter-detector unit 59 , which may have an optical waveguide. The light is absorbed by the mixture and an equally modulated fluorescent light is emitted. The fluorescent light is transmitted from the emitter detector unit 59 to a suitable transmitter (not shown). To measure the fluorescence as a function of light signals, continuous light and / or high-intensity light flashes are / are radiated in addition to the measuring light after a certain period of time via the emitter-detector unit to the mixture in the upper measuring chamber.

The spatial separation of the oxygen and fluorescence measurements solution has the advantage that the measurement is 4 instead of more usual wise 6 minutes of measurement can be limited. For this purpose a high intensity steady light will appear in the lower measuring chamber blasted onto the mixture so that a higher oxygen signal is generated with a more favorable signal-to-noise ratio. With the Fluo Resence measurement in the upper measuring chamber has proven to be favorable proven to use a permanent light of lower intensity so that the relative difference in intensity, based on the additionally flashes of light, remains sufficiently large.

Fig. 3 shows the temporal course of the fluorescence intensity 101 and the oxygen evolution 103 during a playful six-minute measuring cycle in an exemplary embodiment with undivided measuring space according to FIG Irradiated sample. The basic fluorescence 104 is measured, which is a measure of the dark state of the algae. At time t1, a non-saturating flash of light is radiated onto the mixture. The fluorescence intensity 101 shows a sharp and symmetrical tip 105 with a substantially steeper rise on the flank. The curve maximum, the area integral of the curve and the maximum slope in the ascent phase are recorded and taken into account as parameters together with the basic fluorescence in the pollutant analysis. At time t2, a saturating high-intensity flash is emitted onto the mixture and a non-saturating photosynthetic continuous light is switched on. The fluorescence intensity reaches a maximum peak 107 which, based on the basic fluorescence, represents a further (fifth) parameter for the pollutant analysis. After the flash of light t2, only the non-saturating continuous light is radiated onto the mixture over a period of about 3 minutes. The slow decrease in the fluorescence intensity during this photosynthesis exposure time represents a sixth parameter and the limit value, which the fluorescence intensity approaches in the case of photosynthesis light irradiation, represents a seventh parameter for the pollutant analysis. The limit value 109 is also referred to as “steady state value”. Before the steady state is reached, further strong light flashes are emitted onto the mixture at times t3, t4, t5. The relationship between curve maximum and basic fluorescence is taken into account as the eighth parameter in the pollutant analysis.

The development of oxygen 103 is not influenced by the flashes of light at times t1 to t5, since the amounts of energy are too small and the photosynthesis process also reacts too slowly. The development of oxygen is a particularly important measurement when monitoring the influence of pollutants on the physiological processes, since not only the photosysn thesis, but also the dissimilation (breathing) of the algae can be monitored. This happens until time t2 at which the photosynthesis light is switched on. Before t2 the oxygen development slowly drops, since the dissimilation consumes oxygen and not enough (in the case of a completely darkened measuring room) oxygen is not developed as part of the photosynthesis. The drop in oxygen development is a ninth parameter for the subsequent pollutant analysis. The decrease in oxygen generation is a measure of the inhibition of breathing by pollutants. Since different types of pollutants inhibit respiration and photosynthesis to different extents, this parameter, which is based on a different physiological process, can make an important contribution to pollutant analysis.

After t2, switching on the photosynthesis light, takes the oxygen development due to the induced photosyn these too. The increase in oxygen evolution is measured and also taken into account in the pollutant analysis (tenth parameter).

In Fig. 4, the course of the basic fluorescence intensity of the fluorescent light emitted by Rheinwasser is shown as a function of the measurement number of the regularly repeated measurements. In the normal state, the measured values lie within a fluctuation range 111 . The pollutant atrazine introduced at a time of t _s into the Rhine water with a concentration of 1 [µg / l] led to a sudden increase in the basic fluorescence intensity measurements. The measured values in the measurement of the polluted Rhine water lie outside the fluctuation range 111 of the normal state. At this pollutant level, the measurement of the basic fluorescence is sufficient to detect the pollutant. In order to achieve a higher accuracy in the pollutant analysis, however, it makes sense to monitor the oxygen development at the same time and to carry out the pollutant analysis taking into account several of the ten parameters described with reference to FIG. 3.

The quality of the algae suspension is monitored on the basis of the optical density measurement carried out regularly in the algal storage vessel 1 . The suspension is regulated to a certain refractive index by adding nutrient medium, regulating the temperature to about 20 ° C. and adding gaseous carbon dioxide to lower the pH. Under these conditions, the amount of algae doubles approximately every 24 hours. The algae storage vessel is dimensioned so that the regularly removed algae suspension amounts are compensated for by natural growth. An algae suspension can be used for a very long time in this way. Only in the event of a malfunction, if the regulation fails, must the algae suspension be replaced by a new suspension.

There are numerous variations within the scope of the inventive concept lungs compared to the previously described embodiments possible. The mixing vessels can be transported from another who is transported on a possibly interrupted path the; they can be moved from the transport device to a separate one Rinsing station transported and then to take a new one Mixtures are returned to position A. Also can be Any number of mixing vessels rotatable or stationary provided who the. The ratio between the amount of algae suspension and each because the reference water sample used can be the analytical conditions can be easily adapted. For evaluation can any parameter, such as breathing Monitoring parameters of the oxygen consumption before switching on of the photosynthetic light, or any combination of parameters tion can be used.

Claims (21)

1. Process for the pollutant analysis of water samples with algae as indicator means, characterized in that

• a) that an algae suspension ( 9 ) is formed;
• b) that a water sample is mixed with the algae suspension and is brought into contact with the algae sample over a predetermined reaction time;
• c) that the mixture is irradiated with a weak and modulated light signal (measuring light) and with a photosynthetically active, stronger but unmodulated second light signal (continuous light);
• d) that, depending on the continuous light, the course of the oxygen development and the fluorescence signals emitted by the mixture are detected; and
• e) that the measured values of fluorescence ( 101 ) and the oxygen development ( 103 ) as the basis of the pollutant analysis are compared with each other.

2. The method according to claim 1, characterized in that before the process steps c) to e), the mixture initially only is irradiated with measuring light; that depending on the measurement light from the mixture emitted fluorescence signals as a reason fluorescence and the decrease in oxygen evolution little be recorded at least once; and that the basic fluorescence and the decrease in oxygen development as additional measured variables be taken into account in the pollutant analysis. 3. The method according to claim 1 or 2, characterized net that the mixture ho at least once with a flash of light her intensity is irradiated; that the course of this comes from called fluorescence signal is detected and at the Pollutant analysis is taken into account.   4. The method according to claim 3, characterized in that from the course of the emitted depending on the flash of light Fluorescence light the curve maximum, the area integral of the Curve and the maximum slope recorded in the rise phase and be taken into account in the pollutant analysis. 5. The method according to claim 3, characterized in that the mixture with at least one flash of light each before and is irradiated during the photosynthetically active exposure and that the ratios of maximum fluorescence to the bottom fluorescence before and during the photosynthetically active Be clearing as a further influencing factor in pollutant analysis be taken into account. 6. The method according to any one of claims 1 to 5, characterized ge indicates that the process steps b) to e) at least once a measuring cycle is upstream, in which instead of Water sample a reference water sample with known Pollutant content is used and the rest of steps b) to e) to detect the fluorescence and oxygen signals Reference conditions are carried out; that from the relationship of the various signals depends on the pollutant content Reference signal pattern is derived; and that the signal pattern from subsequent pollutant analyzes of water samples the at least one reference pattern for assigning a be agreed pollutant content is compared. 7. The method according to any one of claims 1 to 6, characterized ge indicates that the density of the algae suspension is monitored and is kept substantially constant. 8. The method according to any one of claims 1 to 7, characterized ge indicates that the temperature of the algae suspension is recorded and is regulated to a predetermined value.   9. The method according to any one of claims 1 to 8, characterized in that the two components (algae suspension and reference water or water sample) are pumped into a mixing vessel ( 5 ) with a predetermined ratio, and that the mixing vessel at the end of the measuring cycle emptied and rinsed with water. 10. The method according to any one of claims 1 to 9, characterized in that after the reaction time required for measuring the pollutant-dependent fluorescence signals and the oxygen development, a sample amount is taken from the mixture; that this sample is pumped into a measuring cell ( 29 ); and that after the end of the measuring cycle the measuring cell is emptied and rinsed with water. 11. The method according to claim 10, characterized in that the mixing vessel for filling at a first point (A) statio kept nary, then over the exposure period to a two transferred point (B) and the sample amount there for connection ßendes measurement of the fluorescence and oxygen signals in the Measuring cell is removed. 12. The method according to claim 11, characterized in that a further mixing vessel ( 5 ) is filled at the first point and / or transferred from the filling point to the sampling point during the measurement of the fluorescence and oxygen signals. 13. The method according to any one of claims 1 to 12, characterized in that the algae suspension ( 9 ) and the reference water or water sample are intimately mixed by stirring in the mixing vessel. 14. Device for pollutant analysis of water samples with Al gene as an indicator, characterized in that a Ge water sample line ( 17 ) and an algae suspension feed line ( 11 ) can be coupled to a mixing vessel ( 5 ); that a pro benentnahmevvorrichtung for taking mixture samples from the mixing vessel with a measuring cell ( 29 ) can be connected; and that the measuring cell is assigned at least one light source, an oxygen detection means and a light detector. 15. Apparatus according to claim 14, characterized in that the water sample feed line ( 17 ) and a reference sample feed line ( 19 ) via a valve ( 15 ) are alternatively ver bindable to the mixing vessel. 16. Apparatus according to claim 14 or 15, characterized in that the oxygen detection means as a stick electrode and the Light detector are designed as a photomultiplier. 17. Device according to one of claims 14 to 16, characterized in that the measuring space of the measuring cuvette is divided into two measuring chambers ( 51 , 52 ) which are connected via a capillary ( 53 ), both measuring chambers being a light source which is a measurement Chamber the oxygen detection means ( 57 ) and the other measuring chamber of the light detector are assigned. 18. Device according to one of claims 14 to 17, characterized in that a data processing device ( 33 ) for controlling the functions of the conveyor ( 7 ), the drive of the rotary table ( 3 ) and the measurement signal recording by the oxygen detection means and the light detector and is provided for evaluating the measurement signals. 19. Device according to one of claims 14 to 18, characterized ge indicates that a turntable as (3) carrier at least one  Nes mixing vessel is arranged and designed such that it supports the mixing vessel at the filling point (A) transferred to the sampling point (B) in a circular arc and is also supported there. 20. Device according to one of claims 14 to 19, characterized in that the turntable ( 3 ) is rotatable step by step via a stepper motor. 21. Device according to one of claims 14 to 20, characterized in that the algae suspension supply line ( 11 ) with a temperature-controlled algae suspension storage vessel ( 1 ) is the unit that a device for determining the optical density (refractometer) and a CO₂ supply line having.