Water is a main solvent of many biological experiments and assays. Depending on the source (tap, de-ionized, or ultra-filtered), water purity can differ significantly, which can lead to downstream differences in data reliability and reproducibility. Water purity is particularly important for spectrophotometry applications that make use of transmitted or reflected light to quantify and characterize biomolecules like oligonucleotides and proteins that are typically suspended in liquid media. Spectrophotometry is an important technique in the lab for detecting and characterizing biomolecules and water purification is an essential step to generate accurate and reproducible data from spectrophotometers.
A spectrophotometry primer
A spectrophotometer works by sending a beam of light of a specific wavelength into a cuvette containing the sample of interest suspended in a water-based solvent. Transmission spectrophotometers project light through the sample, and detectors measure the wavelength and quantity of light that pass through. Reflectance spectrophotometers project light onto samples and measure the percentage that is reflected. When transmitted or reflectance light heats a photodetector, it is converted into a current that is electronically amplified, and values such as absorbance or concentration are then generated. Spectrophotometry techniques work with different types of light such as ultraviolet, visible, and infrared light, although the latter is less common.
Spectrophotometry requires highly pure water
Water purity is important for spectrophotometry applications because it is the main solvent in which the biomolecule or sample of interest is being dissolved or suspended. Poor quality water may contain contaminants such as bacteria, endotoxins, and proteases. When water purity is inconsistent, it leads to inaccuracies and poor reproducibility on two levels. First, transmitted or reflected light passing through the cuvettes will differ significantly across different samples. Secondly, the presence of contaminants at different levels may lead to various degrees of sample degradation. Therefore, high purity water is important for reproducibility in spectrophotometry applications.
What to look for in water purification systems
The purpose of water purification systems is to remove contaminants from tap water, including inorganic ions, organics, colloids, gases, bacteria, and proteases. There are a variety of water purification techniques to choose from to ensure high quality water for spectrophotometry applications.
Distillation methods utilize heat to evaporate water and collect the condensate, which aids in removing most contaminants except those with lower boiling points than water. Activated carbon is most effective in removing chloride ions and organic compounds that preferentially bind to carbon. Ultrafiltration makes use of membranes with pores approximately three nm in diameter to remove large particulates, while reverse osmosis makes use of membranes with pores less than one nm for more stringent removal of contaminants like bacteria that are larger than the membrane pore size. Ultraviolet radiation is useful to eliminate potential living organisms like bacteria. Most products utilize two or more techniques to achieve different levels of water purity, and are most effective in removing different types of contaminants.
The first step in choosing a water purification system is to consider the water purity requirement. According to the National Committee for Clinical Laboratory Standards, spectrophotometry and other analytical applications require Type I water.
There are a variety of water purification products on the market including MilliporeSigma’s Milli-Q® Ultrapure Water System, Thermo Fisher Scientific’s Barnstead System, and ELGA’s Veolia Purelab® Quest System to meet the needs of your lab. For a lab with limited space, physical footprint is an important consideration and products that are mobile and do not have to be mounted are more suitable. Scalability should also be considered, as the water purification system has to be able to meet the demand for purified water at a sufficient throughput. For instance, products from Thermo Fisher can provide flow rate from 0.1 L/min (Pacific TII) to 4L/min (Harvey DI+). Some spectrophotometry experiments such as immunoassays to detect proteins in blood samples can be sensitive and, in that case, clinical grade water may even be required, and products such as the ELGA MEDICA® Pro-R would be suitable.