HPLC (A Practical User's Guide) || Advantages and Disadvantages of HPLC

✍ Scribed by McMaster, Marvin C.

• Publisher: John Wiley & Sons, Inc.
• Year: 2006
• Tongue: English
• Weight: 158 KB
• Edition: 2
• Category: Article
• ISBN: 0471754013
• DOI: 10.1002/9780470079096.ch1

✦ Synopsis

The first things we need to understand are how an HPLC system works, its best applications and advantages over other ways of separating compounds, and other techniques that might compliment or even replace it. Is there a faster, easier, cheaper, or more sensitive method of achieving your results? The answer is yes, no, maybe. It really depends on what you are trying to achieve.

HPLC's virtue lies in its versatility! I have used it to separate compounds of molecular weights from 54 to 450,000 Daltons. Amounts of material to be detected can vary from picograms and nanograms (analytical scale) to micrograms and milligrams (semi-preparative scale) to multigrams (preparative scale). There are no requirements for volatile compounds or derivatives. Aqueous samples can be run directly after a simple filtration. Compounds with a wide polarity range can be analyzed in a single run. Thermally labile compounds can be run. I had one customer whose company made explosives for primers. Her first job of the day was to explode samples of the previous day run with a rifle. Her second job was to carry out an HPLC analysis of that day's run.

An HPLC offers a combination of speed, reproducibility, and sensitivity. Typical runs take from 10 to 30 min, but long gradients might consume 1 to 2 hrs. I have seen 15-to 30-sec stat runs on 3-mm columns in hospital laboratories. Retention times on the same column, run to run, should reproduce by 1%. Two columns of the same type from the same manufacturer should give 5% or better retention time reproduction on the same standard set.

While the HPLC can be used in a variety of research and production operations, there are a few places where it really shines. Because it can run