The purity of the major reagent used in all laboratories is often not fully monitored and recorded. This, as Jean-François Pilette reports, means organic contamination is not often determined.
All reagents currently used in laboratories today are delivered with labels indicating the degree of purity and the maximum concentration levels for likely contaminants. Yet, the purity of the major reagent used in all laboratories is often not fully monitored and recorded.
Indeed, laboratory grade water often constitutes more than 99 per cent of the mass of solutions used in laboratory experimentation. Ionic contamination of ultrapure water is generally monitored through resistivity but the degree of water contamination by organic substances is often not determined. This contamination can be reported as the total oxydisable carbon (TOC), a single parameter that takes into account the various organic substances that can be present in pure water and is related to the concentration and the nature of the organic substances.
For instance, acetone (CH3COCH3) has a molecular weight equal to 58 Dalton and contains three carbon atoms with an atomic weight of 12 Dalton each. The carbon ratio in acetone is therefore equal to 36/58 = 0.62 and a 1 ppb acetone solution will display a TOC equal to 0.62 ppb. A similar calculation shows that methanol carbon ratio is equal to 0.375 and that a 2.66 ppb methanol concentration corresponds to 1 ppb TOC. Examples of the concentration required to reach 100 ppb TOC value are given in Table 1 for various organic substances.
The TOC parameter expresses the organic contamination of water containing various organic substances. For instance, a solution containing
133 ppb methanol and 125 ppb formaldehyde will generate 100 ppb TOC. TOC may reach levels of 1000 to 2000 ppb in tap water and up to 5000 ppb in deionised water produced by regenerable ion-exchange resins.
The highly variable organic contamination of water purified by regenerable ion-exchange resins can be attributed to the release of organic substances from the polystyrene/divinylbenzene resin beads matrix after regeneration by strong acid and bases, and to the growth of micro-organisms on old resins used for more than one year.
Organic contaminants in water can interfere in many laboratory applications. In HPLC, LC and ILC, for instance, organic may coat the outside of the solid phase beads and prevent access of the molecules or ions to the exchange sites, reducing the separation resolution; they may also generate a high baseline.
Medical researchers at a national research laboratory developed a sensitive HPLC method for measuring ascorbic acid by electrochemical detection. However, even low TOC (a50 ppb) in the high purity water used to make the mobile phase interfered significantly with the analysis generating a high background and decreasing the analysis sensitivity. Reducing the water TOC <5 ppb increased sensitivity and improved the baseline and quantitation by decreasing background levels. This can be seen in Fig. 1.
In cell culture experimentation, organic substances can affect the cell viability and growth. It is therefore important to make sure that TOC is kept at a low level, and this need has been recognised and expressed in several international norms, such as ASTM, ISO and USP.
Millipore recognised this need early on and developed a compact TOC monitor in collaboration with Anatel, the world leader in TOC measurements. This patented A10 monitor works as follows:
Water flows through the analytical cell, then the solenoid valve closes and the conductivity of the water in the cell is measured, compensated at 25°C. The UV lamp is powered aon' and photocatalytic oxidation begins. The energy provided by the
185 nm UV rays promotes the generation of hydroxyl radicals from the oxygen dissolved in water. These OH radicals oxidise the organic substances in the water sample, ultimately producing carbon dioxide that dissolves in water, yielding bicarbonate ions and protons that cause conductivity increase.
A complex set of algorithm monitors the conductivity variation to ensure complete oxidation of all organic substances and calculates the TOC value from a calibration curve when the final conductivity value is stable and displays the result.
The Millipore A10 TOC monitor is available as a standalone unit that can be linked to a Millipore water purification system, delivering water with resistivity above 5 MW cm.
ENQUIRY No 90
Jean-François Pilette is group product manager, Millipore SA, Saint-Quentin-Yvelines France. www.millipore.com
