HPLC-grade water is something every scientist or researcher gets to use. You have to know several things about it, and there is no way around it.
In this article, find out what HPLC water is, how contaminants affect it, and how to handle it in a correct way.
HPLC-Grade Water
HPLC water is one with low UV absorbance, and it has specific conductivity of 16 to 18 megaohms. So purification systems are obtaining you clean and pure water.
Solvent-rinsed containers are perfect for pure water, and scientists seal it inside. That way, everything is clean and ready for use.
This process is necessary because it avoids contaminations.
Water is pure because of the combination of the following methods:
- Ion exchange,
- Filtration,
- Ultraviolet irradiation,
- Reverse osmosis,
and other methods.
HPLC water must be clean not to interfere with the analysis.
How do Contaminants Influence HPLC Operation?
Several contaminants will have an impact on the results of the HPLC process.
The contaminants are solid particles, gases, leachables, ionic species, organics colloids, etc.
Here is the explanation of each one:
Solid particles
Solid particles in HPLC-grade water can damage pumps and injectors.
Besides, they can shorten the life of a column by increasing the wear and tear on seals, valves, and the pump piston.
Blockage of column frits may raise column backpressure in the long term. That results in a slowing or total cessation of the mobile phase.
Colloids
Colloids may adsorb on the stationary phase forever, lowering column separation efficiency.
Organics
Organic contamination can affect chromatographic separations in a variety of ways.
Organic molecules that build up on the surface can cause resolution loss.
Organic substances in chromatographic separation water can reduce the sensitivity of analytical chromatography. This happens when organic molecules in the eluent water compete with sample molecules.
As a result, the elution process frees fewer sample molecules.
Organics can create extraneous ghost peaks(s).
There may be a lot of organic contamination. The contaminants might act as a new stationary phase. That will cause peak tailing and retention time shifts.
Ionic Species
Ions can also alter chromatographic separations.
Any change in the ionic strength can change the order in which polar molecules separate.
Furthermore, UV-absorbing ions might appear as ghost peaks, complicating data processing.
Leachables
Leached substances from containers are often introduced into high-quality water storage.
Plastic containers can leak organics of HPLC mobile phases or buffers. But, glass containers can introduce ions.
Gases
The development of minute bubbles causes dissolved air in the mobile phase. This happens under the high pressures experienced in the HPLC water system.
These bubbles add to detector noise and column voids.
As a result, you have to use on-line filters to filter the mobile phase and buffers. Use them during preparation and before pumping.
Degassing, whether by on-line or external methods, is critical.
How to Handle Water In a Correct Way
The better the purity of water, the more you need to take care when handling it. That is because contaminations begin when it comes into contact with the air. It’s because water dissolves a wide range of compounds.
If you use commercial water, you should use it before rather than after. It is critical to maintaining ultra-pure water production systems on a regular basis.
You have to limit interaction with air when obtaining commercial or filtered water. Do that by avoiding the formation of bubbles.
Also, pour water straight into the preparation container. Don’t use wash bottles.
Water in wash bottles can get polluted. Also, plasticizers leached from the bottle’s inner walls may be present.
It is of standard knowledge to use freshwater to pre-wash the equipment is necessary.
The risks of using poor-quality water
Almost all trace HPLC studies need the use of large amounts of clean water.
Clean water is a must throughout the whole process:
- Sample preparation
- Pre-treatment
- Solid-phase extraction
- Reagent water for preparing standards
- Blanks to eluent preparation
The purity of the water can be a vital element of the findings produced in these and other applications.
These applications need high-quality water that meets demanding purity requirements. High-purity water is essential for precise, cost-effective, and reliable lab analysis.
The issue is that water includes a wide variety of pollutants. Those pollutants must vanish. Clean water can become polluted in easy ways. Even micro quantities of impurities can damage one of the HPLC steps. That means your result is invalid. It’s compromised.
You have to have a system that will provide you with ultra-pure water for all the research.
Maximizing HPLC Results with Ultra-Pure Water
As Elga Veolia said:
Ultra-pure water is a must for HPLC analysis. The contamination issues explain the “why” part.
If you want to achieve consistent, precise outcomes, using pure water is crucial.
Deionized or distilled water is insufficient. It still contains organic compounds and other contaminants.
Clinical Laboratory Standards Institute’s suggestion is to use Clinical Laboratory Reagent Water. That is a required minimum for general pharma lab work.
But, Clinical Laboratory Reagent Water is often not pure enough for HPLC. HPLC is a sensitive analytical process. Because of that, you have to use type 1 (ultra-pure) water. That is a water of the most incredible purity.
Ultra-pure Water Characteristics
1. Resistivity of 18.2 megohms
2. A total organic carbon (TOC*) of 10 ppb
3. A bacterial count of 1 CFU/ml
*Use Total Organic Carbon (TOC) to measure the organic purity of water.
The water of the most excellent purity is essential for HPLC and trace metal analysis. Besides, it’s needed to produce aqueous mobile phases, buffers, samples, and standard solutions.
As an alternative, laboratories can buy HPLC-grade water from solvent suppliers. Or even establish a water purification system.
For aqueous mobile phase preparation, chromatographers often use HPLC-grade bottled water.
Please keep reading for all information on bottled ultra-pure water and its contaminations.
Bottled Pure Water or In-House Pure Water?
In your lab, you have two alternatives for obtaining water for HPLC:
You have the option of using
1. Bottled HPLC-grade water or
2. An in-house purification system.
You can use bottled HPLC grade water for HPLC eluent and produce standards and blanks. But, depending on bottled water is inferior to utilizing an in-house purifying system.
Costs are more significant when using large amounts of bottled water. Also, a consumer is reliant on the analyses supplied with the bottle, which is based on bulk samples. Besides, you have to have validation on suitability for a specific analysis. Finally, there are significant hazards of contamination after opening the container.
Another study says
When the HPLC-grade bottle is pre-concentrated and eluted, it produces significant peaks. But, these peaks were not present when using at-the-moment created ultrapure water.
Bottled water is generating significant variations in baselines. It’s the same as ghost peaks when used to separate a drug combination.
These issues were not present in the at-the-moment created ultrapure water.
– Bottled HPLC-grade water has a more significant amount of organic pollutants.
– Ultrapure water has less or zero organic pollutants.
Remember that a well-designed in-house purification system guarantees a constant ultrapure water supply. Other than that, the water used for your HPLC testing is clean and of the highest quality.
Can You Drink Deionized Water?
Long story short – yes. You can drink deionized, pure water. But it’s not as simple as that.
Studies from the World Health Organization (WHO) showed exciting results. WHO suggests that people who drink deionized water will urinate more often. That also means that the person will lose more electrolytes from their body.
This is unlikely to be an issue if you get enough vitamins and minerals from the foods you eat on a daily basis.
Although deionized water has a lower pH than ordinary tap water, it is often not acidic. The pH of most carbonated drinks, for example, is much lower than that of DI water.
Most people will not experience any acute health impacts from drinking deionized water.
Submarine crew members consume distilled water on a daily basis. Yet they appear to have no long-term health consequences.
How Does it Taste?
There are two reasons why people don’t drink deionized water:
1. Big costs of acquiring and maintaining a DI (deionized) system, and
2. It doesn’t taste like regular drinking water.
We’ve become accustomed to some degree of impurity in our water. These impurities can be organics or minerals.
When removing everything from the water, many people comment that it tastes “flat” or weird. It’s because humans drink only polluted water and believe that is how water should taste.
Conclusion
Now you know what HPLC-grade water is, how to use it, and what the contaminants are.
Use this knowledge in your field, and always make sure you use the cleanest and purest water. It is a must. You don’t want your chromatographs to be invalid.
