Talk:High-performance liquid chromatography

C18 has been the traditional choice for purification of peptides, but recent work has trended toward the use of C4.

The processes of peptide synthesis involve small molecules and produces by products that are also small molecules. These compounds partition rather than adsorb to the surface of C18. Partitioning moecules will elute much later on C18 than on C4. Larger molecules, such as peptides (>5mer) and proteins, adsorb rather than partition. These by-products with may coelute with peptides on C18 will be widely separated on C4. Peptide elution changes very little.

PRABHUSANKAR,

RESEARCH OFFICER SHASUN DRUGS AND CHEMICALS,CHENNAI

article hard to understand

I read the entry for High performance... and found it hard (as a student who is not familiar with analytical methods) to understand. The terminology was not explained. A diagram (or two) would have been helpful. I am sure that someone who already knows the material might find it well written. ahandel

error

I believe sensitivity has been confused with limit of detection(LOD). Sensitivity is defined as the change in instrument output per unit change in concentration of analyte. LOD is the concentration of analyte that can be detected above the noise with reasonable certainty. Therefore, a sharper peak increases the signal to noise ratio and the limit of detection is decreaseed(better).

Agreeing with the above, i added a clean up tag, maybe someone will fix it...

Effort to improve the article

I have reworked the beginning. I have corrected the error pointed out, added some general easy to understand language at the definition and split the previous language into Isocratic and Gradient HPLC. These two sections could still use some improvement. Perhaps they need to be further split into definition and practical example subsections. On the sensitivity error: this error is an exceptionally common error as to almost be acceptable. I chose not to use LLOD because it is not always relevant; however, signal to noise is always what is being spoken about when people say sensitivity (except when using the term correctly).--Nick Y. 21:23, 5 May 2006 (UTC)

Added ion exchange chromatography section.--Nick Y. 23:00, 5 May 2006 (UTC)

Bead size

Bizarrely in the 10 years I've worked with LC I've never come across the term "Bead" or "bead size" - it has always been referred to as particle or particle size. I wonder if this is a regional thing (I work in a UK university lab)?

138.37.170.200 I presume bead size is a term mainly used in low pressure chromatography, or pellicular methods? I tend to agree that particle size is the usual terminology for modern HPLC.

Well, I have heard the term somewhat frequently from people who actually manufacture stationary phases ( I am one of them). Not sure why. Could be a historic reason. I agree that in all HPLC literature we refer to it as particles.--Xenofonos 00:16, 26 October 2007 (UTC)

UPLC

UPLC is a Waters trademark. These systems are now available from a number of other manufacturers.

True, but it looks likely that UPLC will become the "generic" name - like Hoover, Kleenex etc.. Other comapnies are calling them different things but usually with the word Ultra, or Ultra-high pressure/performance LC - which I am hearing more and more as "UPLC". --Daeve 22:16, 17 July 2006 (UTC)

@Daeve: I don't agree. It's not a good idea using trademarks and brandnames to describe general methods and techniques. This applies to an encyclopedia as well as to scientific publications. Everytime naming it "UPLC", would mean an infringement of Waters' rights and would require to mention in any publication that it is a trademark of the company. In analogy to "ultra high temperature" (UHT) compared to "high temperature" (HT) and "ultra high frequency" (UHF), compared to "high frequency" (HF), the term "ultra high performance liquid chromatography" and its acronym (UHPLC) would be the correct one compared to "high performance liquid chromatography" (HPLC) and should be used instead. "UHPLC" cannot be easily turned into a trademark by any company, because it's systematically deducted and lacking any uniqueness. --Chemist at work 04:20, 3 February 2007 (UTC)

Another possible error (?)

In the Reversed phase chromatography section one can read the following sentence:

... Eluent time is increased by addition of less polar solvent.

Although I'm not sure what Eluent time means, I think this deduction is wrong. In a RPLC the stationary phase is non-polar. Addition of a less polar solvent will make the solvent compete for the stationary phase with the analyte, thus freeing it more quickly, which, in turn, decreases the time needed for a substance to elute. jοτομικρόν (talk, email) 23:00, 5 November 2006 (UTC)

Corrected. There were a series of edits a while back that suffered from imprecise and somewhat confusing language. They did however make some valid point as is the case here, which is why they were not reverted. Adding polar solvent to the mobile phase does indeed increase retention time. But "less polar solvent...increasing elutant time" is definately wrong however you slice it but seems very close to correct on first look. E.g. change the unusual phrasing of "retention time" as "elution time" to the more common phrase "elution rate". Anyway I have ment to clean the whole thing up but was hoping the contributor of these additions would do so himself. Feel free to improve the article as you see fit. Someday I will give it a thorough cleaning but some other good editing would be appreciated before I tackle it.--Nick Y. 18:25, 6 November 2006 (UTC)

For some reason I went ahead and cleaned up the article. Any feed back is good though. --Nick Y. 20:35, 7 November 2006 (UTC)

"high performance"

The article states that the term "high pressure liquid chromatography" has been deprecated in favor of "high performance liquid chromatography". The word "performance" was introduced as part of an advertising campaign for high-pressure liquid chromatographs, where the advertisers said that "the P in HPLC stands for performance". Evidently, the scientists who bought the machines were naive enough to believe them. But if the P stands for "performance", what the heck is MPLC? "Medium performance liquid chromatography"? Please, let's reject this advertising-induced nonsense and restore the original, meaningful term. —The preceding unsigned comment was added by Rbgros1 (talk • contribs) 14:05, 31 January 2007 (UTC).

My understanding is different. Although pressure and performance are frequently interrelated performance is not fundamentally connected to pressure. The most obvious example is that performance will drop dramatically at too high of a pressure for the system when the kinetics break down. Pressure drives optimal flow rate and generally increases with surface area which is the first order determinant of performance. This is the reason for the change to performance somewhere around twenty years ago. Pressure is still used by some and is not entirely unacceptable; however it is not the primary explanation of the acronym in the IUPAC gold book [1]. We should follow the IUPAC convention. It is probably a good idea to revisit our naming convention however since there is a dash between high and performance.--Nick Y. 19:42, 31 January 2007 (UTC)

I am with Nick on that one. The introduction of smaller particles was the driving factor in improving performance. The down side was the need for much higher pressures. The pressure is a result of the improved performance and not vice versa.--Xenofonos 17:58, 25 October 2007 (UTC)

It should be pointed out the what the Wikipedia audience thinks it *should* be called is completely irrelevant. Names need not be descriptive. One can argue that the "Democratic People's Republic of Korea" (the official name of North Korea) is in truth neither democratic, nor a republic, nor "of the people" - yet it still gets the moniker "Democratic People's Republic of Korea". Likewise, what WP users say HPLC "should" stand for is irrelevant - what matters is what it *is* called, and what term is accepted as "official" by experts in the field. The IUPAC link that Nick gives shows that "Performance" is the preferred term, but "Pressure" seems to be given as an acceptable alternative.

The OP brings up a good point, though, which isn't covered in the current article: What's the history of the Performance/Pressure split? Where did it come from? Which was the "original" term? Why does the split persist? Is there any commercial/administrative forces which were at work vying for one side or the other? Who were the players in standardizing on "performance" as the preferred term, and why? It is a fact that both terms are/have been used, and the reason for the differing terms is an encyclopedic topic that should be covered, if appropriate references can be found. -- 17:24, 19 December 2007 (UTC)

I agree that the split in terms is an encyclopedic topic which is worthy of coverage. I think you agree that we should use "Performance" as it is the most common today and recommended by harmonizing bodies. To my understanding pressure came first. I don't think the work of researching the evolution of the name has been done and would require looking at historical usage in the primary literature. That would be bordering on original research but not really?--Nick Y. (talk) 23:34, 19 December 2007 (UTC)

I reverted and edit by an anonymous user who removed the "high-pressure liquid chromatography" from the first sentence of the article. Googling "high-pressure liquid chromatography" yields several hits, and although high performance is probably more common, it seems as though the consensus was that both terms should be discussed or noted at the very least. Pdcook (talk) 15:49, 28 September 2009 (UTC)

Manufacturers sections

Please keep this section a resource deserving the name "manufacturers". Thus, any company not designing and manufacturing HPLC systems or instruments, should not be named in the "chromatographs" section and companies not manufacturing HPLC columns and supplies should not be listed in the "columns and accessories" section. Otherwise this list will become as irrelevant as many other lists published by magazines, buyers guides and so on. The instrument manufacturers list on "Links for Chemists (part of the WWW Virtual Library)" maintained by the Liverpool University (UK) is a relevant one, but not limited to chromatography. Letters a-j: [2], letters k-z: [3].--Chemist at work 13:30, 3 February 2007 (UTC)

I don't see why an encyclopedia needs a buyers guide of manufacturers at all. This type of section begs for advertising like contributions, so I propose we delete them.Heckendorf 21:17, 14 March 2007 (UTC)

Although I see value to these links, I also think that the section is a little out of hand and turning into somewhat of a spamvertisement section. I might suggest that we turn all of them into internal links only, and then only those with articles of at least stub size that is properly referenced and claims and demonstrates notability as a manufacturer. I may proceed with this if I don't hear any other suggestions.--Nick Y. 17:18, 21 March 2007 (UTC)

I'm a Wiki writer newbie and want to agree to Nick Y.'s opinion to just keep internal links to real manufacturers, not resellers. What means stub in this respect? -- guest —The preceding unsigned comment was added by 84.191.232.11 (talk) 10:37, 11 May 2007 (UTC).

I added HP (Hewlett Packard) to the list of manufacturers as they do make HPLC products, including the one used in my lab.--76.105.4.141 (talk) 02:40, 10 November 2008 (UTC)

But they don't - they used to, but they spun their instrument operations (including HPLC stuff) off as Agilent in 1999. Agilent is already listed. -- Jaeger5432 | Talk 16:53, 10 November 2008 (UTC)

I added Advanced Materials Technology, Inc. they are the inventors of Fused-Core Technology and manufacture the Halo brand columns. —Preceding unsigned comment added by Jayjay02 (talk • contribs) 22:21, 22 December 2009 (UTC)

I added Knauer, a HPLC equipment manufacturer based in Germany. Will write up an article for them later.--89.212.75.6 (talk) 22:50, 2 January 2010 (UTC)

Principle

The first paragraph previously started with "In isocratic HPLC..." but has been changed to "The basic operating principle of HPLC is...". Although the first sentence is clearly generic, other sentences in the first paragraph describe isocratic HPLC. Specifically the sample being retarded as it traverses the column is very isocratic. In gradient elution one might say that the analyte is retained by the column until the solvent composition changes to bring the analyte into solution and traverse the column freely.--Nick Y. 17:12, 2 August 2007 (UTC)

Added infobox

I am testing a new infobox {{Infobox chemical analysis}}. Comments? --Kkmurray 19:15, 15 August 2007 (UTC)

RPC

I don't agree that RPC is a commonly used abbreviation for reversed-phase HPLC. —Preceding unsigned comment added by 82.45.237.9 (talk) 14:56, August 27, 2007 (UTC)

I guess if I saw the abbreviation RPC within the context of HPLC, I would assume it had something to do with reversed phase chromatography. However, if someone said merely I do a lot of RPC at work, I'd have no idea what they meant unless given a context. It's probably best to say reversed-phase HPLC. Pdcook (talk) 19:25, 28 September 2009 (UTC)

Deleted sentence (solvent polarity and retention time)

I deleted the sentence "An investigator can also decrease retention time by adding a polar solvent to the mobile phase, or increase retention time by adding a more hydrophobic solvent." from the section on Reversed Phase chromatography. This claim or its reverse seems to be controversial, eg see this edit, the section "Another possible error (?)" above, and in particular this archived thread at Wikiproject Chemistry. Rather than risk error, it seems safest to omit the sentence entirely, at least until we can cite a clear, authoritative source. Adrian J. Hunter^{(talk•contribs)} 07:22, 10 February 2009 (UTC)

Not a bad move. However, it is not at all controversial and has been established for half a century. Simply either a bad edit from a mistaken person or an intentional effort at subtle vandalism. The previous version is correct and essential to how RP-HPLC works.--Nick Y. (talk) 18:03, 10 February 2009 (UTC)

I have reintroduced the information since it is fundamental to the technology but explained and clarified the statement to avoid confusion and future mistaken editing. --Nick Y. (talk) 18:13, 10 February 2009 (UTC)

selectivity in gradient elution, is this right?

The article contains the following mysterious statement:

"In isocratic elution, the selectivity does not change if the column dimensions (length and inner diameter) change - that is, the peaks elute in the same order. In gradient elution, the elution order may change as the dimensions or flow rate change.[citation needed]"

I am struggling to think of a situation where I could reverse peak-order in a gradient elution merely by changing the flow rate. If no one else can think of such a situation, and no one can cite anything, perhaps this line should be removed? 149.155.96.5 (talk) 16:39, 28 September 2009 (UTC)

I also think this statement should be removed; however, I do think that it is a nominally correct statement. It is not exactly an important or general statement however. I believe the statement is addressing issues with scaling that are not so much theoretical as practical. Scaling generally results in a change in the gradient shape, regardless of programming. I.e. no pump delivers perfect gradients that turn on a dime and the relationship between what you ask for and what you get does change with scale. Dwell times increase, linear diffusion changes etc. It is possible to get a reversal of retention times because of such effects, particularly when you are working with a method that is not purely homogeneous in retention mechanism and such peaks are early in the gradient or at a sharp turn. Here's and example [4] which would make a suitable reference. A more general statement about the scaling of gradient based methods would be better.--Nick Y. (talk) 18:51, 28 September 2009 (UTC)