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Sampling and preparation techniques of wastewater samples

Representative sampling of wastewater streams is decisive for correct modelling of wastewater treatment processes. While in laboratories usually high efforts are made to execute chemical analyses of wastewater samples with high accuracy, wastewater sampling is sometimes carried out by people who are not trained in sampling. Thus, experts assume that errors in wastewater analyses caused by mistakes during sampling are several orders of magnitude higher than by analytical errors in the chemical laboratory (Sommer, 1995).



Different kinds of sampling are possible: grab (or catch) samples and composite samples (a mixture of grab samples collected at the same sampling point at different times) can be taken. Both kinds of sampling can either be carried out manually or automatically. Automatic samplers are being used increasingly. They are effective and reliable and can significantly increase the frequency of sampling. Especially for composite samples taken during long periods (days, weeks), automatic samplers are convenient and help to save manpower. An example is outlined in figure 10. With this type a variable number of constant volumes of single samples can be combined to composite samples. At desired times, which can be programmed by means of a control unit, the vacuum pump A is switched on automatically until the sample reaches the level indicator sensor C in the dosing vessel D. Then the level indicator gives a signal to switch off the vacuum pump. The sample flows back through the tube B until the level in the vessel D is reached which is given by the length the tube B is inserted into vessel D (adjustable). Then the valve E is opened automatically and the sample is flowing into the desired bottle H (also programmed via control unit of the automatic sampler). In many parts of the world, composite samples representing a 24-h period are considered standard. In Germany, for many control parameters composite samples consisting of five grab samples collected within a two hours period are according to regulations.

Figure 10: Scheme of an automatic sampler taking samples by means of vacuum; A: vacuum pump; B: pipe for sample transport from wastewater stream; C: level indicating sensor; D: dosing vessel; E: valve; F: cock which is moved by steps by a small motor in order to select sample containers H for different composite samples; G: channels for distribution of samples to sample bottles H; Gulyas (1999).



Figure 11: Different types of sampling using automatic samplers; A: volume flow of the wastewater stream which has to be characterized; B and C: continuous sampling modes; D, E, F: discontinuous sampling modes; for details see text; Gulyas (1999).



Figure 11 shows the volume flow of a wastewater stream (A) and different types of composite samples (B to F). For continuous sampling, pumps are used either with a continuous flow (time-continuous sampling, A) or with a flow which is adapted to the flow of the wastewater stream (flow-continuous sampling, B). In the schemes D to F (discontinuous sampling) at particular intervals grab samples are taken (being combined to form composite samples) in different ways: In scheme D the frequency of taking grab samples is constant, but the volume of each grab sample is adapted to the volume flow of the wastewater stream (flow-proportional sampling); in sampling mode E frequency as well as volume of grab samples are constant (time-proportional sampling); in scheme F the volume of each single sample is constant, but the frequency is controled by the flow of the wastewater (high sampling frequency during high wastewater flows, low sampling frequency during low wastewater flows, volume-proportional sampling). For sampling modes C, D, and F a flow meter is needed to determine the wastewater flow. The flow meter must be able to transfer its signals to the automatic sampler in order to control flow (mode C), volume (mode D), or frequency (mode F) of grab samples. The automatic sampler drawn schematically in figure 10 can only be applied for sampling modes E and F, because the volume of each grab sample cannot be varied during the sampling period and because it is a device for discontinuous sampling. It has to be noted, that automatic samplers have to be cleaned before and after sampling campaigns. Careful maintenance is a prerequisite for appropriate function.



Table 4 shows different conditions of wastewater flow and concentrations of wastewater constituents and the suitable sampling mode to yield representative samples.

Table 4: Selection of appropriate sampling mode (from several possibilities given in figure 9) yielding representative samples with different conditions concerning wastewater flow and concentration of waste­water constituents.



Besides sampling times, also the location of sampling is very important in order to obtain representative samples. It is recommended to sample below the surface of the wastewater in order to avoid contamination of the sample with materials with a lower density than water which are eventually enriched at the surface but not representative for the bulk wastewater stream. It should also be avoided to take samples directly at the walls or at the floor of e.g. wastewater channels or tanks, because in this case solid substances fixed at the walls (growing biofilms) will be added to the sample although they are not representative for the wastewater stream.



If suspended wastewater constituents are to be analysed care has to be taken, because particle size is easily affected by sampling measures. By transferring mechanical energy to samples (e.g. by shaking or by pumping) two effects can occur with the particles: flocculation or destruction of flocs leading to a greater particle number but lower particle diameters. This can lead to artefacts when particle properties are measured which correspond to particle size: e.g. determination of settleable solids or particle size distribution. For the determination of these parameters, samples taken manually and with care are favorable. Moreover, these parameters should be measured immediately after sampling, before flocculation or floc destruction has occurred.

Figure 12: Analytical errors caused by unsuitable sample containers; A: unsuitable container materials; B: impurities; C: leaky seals; D: unsuitable gaskets; Gulyas (1999).



There are also particular requirements concerning the containers for collection and storage of samples (see figure 12.): First of all, sample bottles have to be clean (to prevent sample contamination) and dry (in order to prevent sample dilution). Cleaning should be performed in the laboratory prior to sampling following standard cleaning procedures. Besides contamination, impurities (B in figure 12.) can also cause adsorption of analytes (i.e. the substances which are to be analyzed in the sample) leading to analytical results below the true concentrations of the analytes in the wastewater. The material (A in figure 12.) of the sample container may also affect the results of analyses: analytes can be adsorbed by unsuitable container materials, sometimes they can pervaporate through the material (e.g. halogenated organic solvents in wastewaters can permeate the walls of polyethylene bottles resulting in too low results or they can contaminate the sample if they diffuse from the air in the laboratory into the bottle), and some constituents of container walls (e.g. plasticizers in PVC bottles) may migrate into the sample leading to sample contamination (resulting in too high results for organic sum parameters like total organic carbon). When sample containers are not tightly sealed (C in figure 12) either volatile analytes can leave the bottle. On the other hand, a laboratory Environment bearing relatively high concentrations of volatiles may cause contamination of the sample. Long term storage of the samples with leaky seals may result in water evaporation from the sample bottle leading to increased concentrations of all non-volatile analytes. Unsuitable gaskets of seals (D in figure 12) may also cause problems either with contamination of the sample (e.g. organic materials) or with adsorption of analytes. Another aspect is the transparency of the container material: Wastewater constituents which are susceptible to photoreactions require bottles which prevent the sample from light (using brown glass of Teflon bottles). Generally, samples should not be allowed to stand in the light, but always stored in the dark.



As most wastewaters are not at all sterile, it can be assumed that biochemical processes (which are desired in biological treatment stages) will not stop when the wastewater sample is transferred to a bottle. Examples for such processes are nitrification (oxidation of ammonia to nitrite or nitrate if still oxygen is dissolved in the sample resulting in too low ammonia and too high nitrite, or nitrate concentrations determined), denitrification (reduction of nitrate or nitrite to nitrogen gas resulting in too low nitrate or nitrite concentrations determined) or oxidation of organic wastewater constituents (resulting in errors of organic sum parameters analyzed in the sample). To prevent biochemical reactions, the microorganisms in the wastewater sample have to be killed or at least their metabolic activity has to be inhibited. This can be obtained by preserving the samples. Preservation can be realized by the addition of substances which are inhibiting or toxic towards microorganisms (acids, bases, mercury salts, azide) or by reducing the temperature (storage of samples in refrigerators or freezers), thus decelerating biochemical reactions.



For different analysts there might be different optimum preservation procedures. For additional details for preservation of samples see appendix in page 40. This means that sometimes a sample has to be divided and the sample parts must be preserved in different ways each adapted to the parameters that are aimed to be analyzed. In the past, there have been attempts to find optimum preservation methods for several wastewater routine parameters. The "Working Party on Stabilization of Samples from the Hydrochemistry Team of the German Chemists Association" (1981) analyzed primary clarifier effluent of a municipal wastewater treatment plant for different parameters after different storing periods and applying different preservation methods (see table 5) and compared the analyses to those executed with the same samples directly after sampling. By this, they could determine the periods of stability for several parameters under different preservation conditions (storing at room temperature without any addition of preservatives and with addition of acid or base or mercury chloride and storing in a freezer).



In the "Standard Methods" also effective preservation methods are given for several parameters (Greenberg et al. 1985). It has to be mentioned that freezing is not a good preservation method for samples to be analyzed for suspended solids. It can be observed, that even in filtered samples after freezing and thawing solids are generated. These solids are a consequence of flocculation of colloidals (dissolved macromolecules like humic substances). During the freezing process more or less pure ice is formed at the walls of the sample bottle leading to increasing concentrations of dissolved wastewater constituents in the remaining solution. The more ice freezes the more concentrated the residual solution will become. Macromolecules come so close to each other that some of them "stick" together and will no longer be dissolved because of the huge size of the agglomerates. When the whole sample is frozen, these solids are also frozen in the ice, but after thawing they are not re-dissolved, but stay solids. Another fact that has to be remembered when applying freezing as preservation is to avoid glass bottles because they can be destroyed during freezing the aqueous sample and can no longer protect the sample against contamination or evaporation of water molecules.



Parameter

Preservation Method

Period of stability of parameter [d]

Oxidation with KmnO4

no preservation

-18 to -22°C

acidified (pH 2)

alkaline (pH 12)

HgCl2

32

16

8

8

COD

no preservation

-18 to -22°C

acidified

alkaline

32

TOC

no preservation

-18 to -22°C

acidified

alkaline

32

2

8

BOD

no preservation

-18 to -22°C

acidified

alkaline

32

4

8

ammonia

no preservation

-18 to -22°C

acidified

alkaline

HgCl2

16

32

32

nitrate

no preservation

-18 to -22°C

acidified

alkaline

HgCl2

8

1

4

sulfate

no preservation

HgClv

32

anionic surfactants

no preservation

acidified

HgCl2

32

Table 5: Recommendations of the "Working Party on Stabilization of Samples from the Hydrochemistry Team of the German Chemists Association" (1981) for preservation of primary clarifier effluent for analyzing different parameters



Efficiency of particular preservation methods strongly depends on concentration of microorganisms in the samples. Therefore, preservation recommendations given in the literature may not always be suitable and applied preservation methods should be verified with samples routineously collected. When using chemical preservatives like acids etc. one should take care, that certain analytes can no longer be determined in a sample preserved in such a way. It is impossible to measure e.g. nitrate or total nitrogen in a sample that had been preserved by addition of nitric acid. Chloride cannot be determined if hydrochloric acid had been used as a preservative.



A couple of parameters are recalcitrant against preservation and have to be measured immediately after sampling. Such parameters are given in table 6.



Parameter

Measures to be taken

turbidity

immediate inspection and documentation; analytical quantification should be carried out on the same day

settleable solids

immediate analysis using Imhoff cone

suspended solids

filtration and gravimetric analysis must be performed as soon as possible

colour

immediate inspection and documentation

odor

immediate check and documentation

concentration of dissolved oxygen

analysis with oxygen probe

pH

analysis with pH probe

conductivity

analysis with conductivity probe

nitrite

transport samples as fast as possible to laboratory for analysis; reflectometric analysis at sampling location

temperature

directe determination in the wastewater

Table 6: Parameters which cannot be stabilized by sample preservation and have to be measured immediately after sampling at the sampling location or directly in the wastewater



Each step of handling the samples has to be documented in the sampling protocol which should also contain the sample designation (which has to be marked also on the sample container), date and day time of sampling, sampling location, name of person collecting the samples, purpose of sampling, mode of sampling (grab or composite sample etc.), results of measurements performed at the sampling site, sample preparation measures (e.g. sedimentation of sample), preservation procedure(s), sample storing conditions until delivery to laboratory, comments upon reference samples simultaneously collected, comments about subsequent changes occurring in the sample, comments about deviations from routineously performed sampling (e.g. application of another automatic sampler, more frequent transfers of samples to other bottles than usually done), observations at sampling site (weather, wastewater irregularities as foam, bulking sludge, odor etc.), comments about irregularities observed on the sampling site (e.g. construction operations within a treatment plant etc.). Sampling documentation forms can serve as check lists.



For further analyses in the laboratory, samples must be transported as soon as possible to the laboratory. For keeping the samples unchanged during the transport, the sample containers should be tightly sealed, kept cool (e.g. using a cooling bag - which should be exclusively used for sampling but not for food transport for safety reasons) and dark. In vehicles used for sample transport, samples must be protected against being tilt over. If samples are shipped by mail or express services, by railway, ship or aeroplane, special safety measurements have to be taken. The bottles must be sealed absolutely tight and protected against shock in order to avoid leakages of the sample bottles.



It is clear that sampling of wastewater (and also of other media) has to be carefully prepared (providing sampling equipment like suitable sample bottles in sufficient number etc.). There must be a good communication between sampling staff and the analytical laboratory concerning number of samples, parameters which must be analyzed, time of delivery of samples to the laboratory, because the laboratory has to organize the enforcement of the analyses as well as to provide storing space in refrigerators or freezers. The samples as well as the sampling protocols have to be received by the laboratory staff in a responsible manner because of registration and eventual transfer of some samples to other laboratories for special analyses.



Working safety has to be obeyed not only in laboratories, but also during sampling (e.g. marking samples with symbols for hazardous materials if harmful preservatives like concentrated acids or bases or even toxic materials like HgCl2 are added to samples).



Another step which is often performed prior to sample analyses is sample homogenization. This is necessary when samples which contain solids are divided. Measures have to be taken that the divided samples are identical with the original sample. This is not possible if a sample contains e.g. settleable solids and is not sufficiently agitated during sample division. Then the solids will settle, the sample is no longer homogeneous and the sample is divided into one part being poor in solids and the other one being more concentrated in solids than the original sample. This can be avoided by transferring an aliquot from the stirred sample. Sometimes high speed stirring devices have to be used in order to keep the sample in a homogeneous state during sample division, see also the presentation (slides of lesson A1).

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