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The equilibrium process can be influenced by several factors which include adjustment of pH to prevent ionization of acids or bases, by the formation of ion-pairs with ionizable analytes, by the formation of hydrophobic complexes with metal ions, or by adding neutral salts to the aqueous phase to reduce the solubility of the analyte (also known as ‘salting out’).
In discontinuous extraction, the most common approach uses a separating funnel (Figure 8.1). In this case, the aqueous sample (1 l, at a specified pH) is introduced into a large separating funnel (2 l capacity with a ‘Teflon’ stopcock) and then 60 ml of a suitable organic solvent, e.g. dichloromethane, is added. The vessel is then sealed with a stopper, and shaken vigorously, either manually or mechanically, for 1 -2 min. This shaking process allows thorough interspersion between the two immiscible solvents, thereby maximizing the contact between the two solvent phases and hence assisting mass transfer, and thus allowing efficient partitioning to occur. It is necessary to periodically vent the excess pressure generated during this shaking process.
What do you think would happen if you did not release the pressure in the separating funnel?
Figure 8.1 A separating funnel used for (discontinuous) liquid-liquid extraction. From Dean, J. R., Extraction Methods for Environmental Analysis, Copyright 1998. © John Wiley & Sons Limited. Reproduced with permission.
The simplest effect would be to ‘pop off the stopper (the path of least resistance). More drastic effects could result if the stopper could not be removed by the excess gas built up inside the separating funnel. This is why in the laboratory it is necessary to wear both a laboratory coat and safety glasses to guard against the unexpected.
After a suitable resting period (10 min), the organic solvent is run off and retained in a collection flask. Fresh organic solvent is then added to the separating funnel and the process repeated again. This should be carried out at least three times in total. The three organic extracts should be combined, either ready for direct analysis or pre-concentration (see Chapter 10), with the exact requirement depending upon the level of contamination.
In some cases the kinetics of the extraction can be slow, such that the equilibrium of the analyte between the aqueous and organic phases is poor, i.e. Kd is very small (see Box 6.1 earlier); if the sample is large, then continuous liquid-liquid extraction can be used. In this situation, fresh organic solvent is boiled, condensed and allocated to percolate repetitively through the analyte-containing aqueous sample. Two common versions of continuous liquid extractors are available, using either lighter-than or heavier-than-water organic solvents (Figure 8.2). Extractions usually take several hours, but do provide concentration factors up to 105 (an essential requirement for trace analysis). Obviously, several systems
144 Methods for Environmental Trace Analysis
Figure 8.2 A typical system used for continuous liquid-liquid extraction, employing an organic solvent which is heavier than water. From Dean, J. R., Extraction Methods for Environmental Analysis, Copyright 1998. © John Wiley & Sons Limited. Reproduced with permission.
can be operated unattended and in series, thus allowing multiple samples to be extracted. Typically, a 1 l sample, pH-adjusted if necessary, is added to the continuous extractor. Then, organic solvent, e.g. dichloromethane (in the case of a system in which the solvent has a greater density than the sample), of volume 300-500 ml is added to the distilling flask together with several boiling
chips. The solvent is then boiled, in this case with a water bath, and the extraction process allowed to occur for between 18-24 h. After completion of the extraction process, and sufficient cooling time, the boiling flask is detached and solvent evaporation can then occur (see Chapter 10).
Unfortunately, as with most things, liquid-liquid extraction can suffer from some problems which affect its efficiency.
A common problem in LLE is the formation of emulsions, particularly for samples that contain surfactants or fatty materials. What do you think can be done to reduce or eliminate this problem?
The emulsion can often be broken up by using one of the following approaches, i.e. centrifugation, filtration through a glass wool plug, refrigeration, salting out, or the addition of a small amount of a different organic solvent.
In addition, the rate of extraction may be different for the same analyte depending on the nature of the sample matrix. Obviously, as in all analyses the problem of controlling the level of contamination is crucial. It is essential to use the highest purity solvents (as any subsequent concentration may also concentrate the impurity as well as the analyte of interest) and to wash all associated glassware thoroughly. As well as contamination, care should also be exercised to minimize analyte losses due to adsorption on glass containers. A typical procedure for LLE is described in Figure 8.3.