Questions tagged [chromatography]

collective term for a set of laboratory techniques for the separation of mixtures

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Decomposable solvent identification in GC-FID

How can a gas chromatography with flame ionization detection (GC-FID) be used to identify a liquid solvent that decomposes at its boiling point like DMSO? In a solution with other higher boiling ...
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Activating alumina before column chromatography

Recently I synthesized some amine compounds, and because it showed better separation in the neutral alumina TLC plates than silica plates, I am planning to purify the reaction mixture using neutral ...
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How much analyte should I use for paper chromatography?

I am looking into using paper chromatography to distinguish between standard drugs and counterfeit drugs for my high school research project. I have decided on using aspirin as the "standard"...
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Solvent for Terephthalic Acid TLC

We're trying to do thin layer chromatography to assess the purity of Terephthalic Acid. So far we have tried DMF, and Water of different pH values. Does anybody know a better solvent or combination?
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Triethylamine is detected by ninhydrin during TLC

Continued from this question. I purchased new triethylamine from Sigma Aldrich, spotted freshly opened triethylamine to the TLC plate, dipped into ninhydrin solution, and heated it. Again the purple ...
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Does ninhydrin staining detect triethylamine?

My reaction mixture has amine compounds, so it shows tailing in bare silica plates. Therefore I decided to add small amount of triethylamine in the eluent. Because my amine compound is not detected by ...
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Corruption of TLC plates or solvent

I have been conducting TLC experiments, and sometimes I observed some lines when I use mixture of eluents. And I found that some lines are observed after I develop the TLC plates with some eluents. ...
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Fragmentation behavior of spots in TLC

I tested multiple development using eluent condition hexane + 1% of ethyl acetate. Then I observed kind of fragmentation of spot. I'm not sure, but I guess that successive spots are from the skyblue ...
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Ion Exchange chromatography [closed]

Suppose that Anion exchanger is used in the stationary phase normally -ve charge A.A will be eluted first as it will not attach but my question is does the zwitter ion(have net charge 0 will be the ...
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Using neutral or basic alumina in column chromatography for purification of amines

I am conducting a synthesis involving amines, and it is found that some spots still hardly moves from the starting point in spite of the dichloromethane 9:methanol 1 eluent condition. Addition of ...
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Can I expect the same (or very close) peak areas from herbal extracts prepared with the same plant/solvent ratio?

I prepared an extract by using 100 mg of plant and 2 mL of ethanol, filtered and injected 20 μl into the HPLC system equipped with Waters® XSelect (150 mm × 4.6 mm, 5 μm) column. If I prepare an ...
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Can I scrape off compounds from normal TLC like in prep TLC for MS analysis?

As far as I know, the limit of detection of mass spectrometry is very low compared to other analysis. In principle, so I think I can scrape off some compounds from normal TLC plate after development ...
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TLC monitoring of Buchwald-Hartwig coupling reaction using ninhydrin stain

I am conducting a synthesis involving Buchwald-Hartwig coupling reaction. The reagents are 2,3,5-tribromothiophene, 1-aza-18-crown-6, NaOtBu as base, $\ce{Pd_2(dba)_3}$ as palladium source, DavePhos ...
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Separation of very nonpolar mixtures using column chromatography

I am conducting a synthesis using Buchwald-Hartwig coupling. Below is the TLC result after nearly all consumption of limiting starting material. Th is 2,3,5-tribromothiophene, which is the limiting ...
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How can I determine the purity of an oligonucleotide solution?

After desalting an oligonucleotide via gel chromatography, I am left with (presumably) an oligonucleotide substance suspended in a buffer. What is the simplest way to verify the oligonucleotide's ...
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Why can I extract all metabolites from blood plasma with one solvent, but to extract only fatty acids I must use many solvents?

I want to do a metabolite study using GC- without derivatization and UPLC-MS (RP and NP) on samples of plasma and urine. My hypothesis is agnostic about which metabolite will correlate with the ...
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Interpretation of permanganate-stained TLC spots

When I stain TLC spots with $\ce{KMnO4}$, I found that some spots appear differently as time goes by. Here is the example: The leftmost one is the plate right after staining, and the others are ...
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chromatographic interactions with methanol, acidic or basic?

How can I describe the interaction between methanol and thiophene in a chromatographic separation in an acid media? I'm confused with the different concepts: Methanol present a hydrogen acceptor ...
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Dark smudge at the TLC solvent front

I have recently started using TLC in my graduate work to test for the cleavage of a cyclic oligo-adenylate in an enzymatic reaction. The sample I'm spotting on my plate is in an aqueous solution ...
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Has anyone added dyes to a mixture about to separated by chromatography in order to visualize where the desired product lies?

Usually in preparative chromatography one needs to know when the desired substance to be separated is coming down the pike. Often this required some sophisticated instrumentation. But, it occurred to ...
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Gas chromatography of samples containg water

I have read a few articles where it is stated that samples containing water are detrimental for a GC experimental results and equipment. For example, in the paper Identification of essential oil ...
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Why is Ion Exchange Chromatography done in a column?

It seems like the idea of Ion Exchange Chromatography is to get a maximum amount of exposure of your gel to your solution. It seems like overhead stirring and a gel bed would be more likely to be ...
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Optimal lipophilic solvent for washing PEEK-lined reversed-phase HPLC columns

What would be the reasonable strategy for washing a reversed-phase (RP) octadecylsilyl (ODS) C18 HPLC columns from lipids and other non-polar residues? The columns are used for biochemical analysis of ...
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What is the function of thin layer in thin layer chromatography?

I just learned about the thin layer chromatography, which I think is basically the same as the paper chromatography. I am wondering what is the function of the thin layer? I mean, what's special about ...
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Impact of flow rate on peak areas in Mass Spectrometry

As a follow up to the discussion in comments: it's now clear that UV peaks will have larger area if flow rate is decreased, because the same portion of molecules are measured again and again. With ...
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Why are cations detected in capillary gel electrophoresis with a positive applied voltage and a coated fused silica capillary?

In capillary gel electrophoresis, when a positive voltage is applied to a coated fused silica capillary, why are positively charged ions (among positive, negative and neutral) detected first? What is ...
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Does triethylamine affect all materials in TLC?

In TLC, I heard that if I treat acid-sensitive materials such as amines, then I can add some triethylamine. The principle of TLC is interaction between materials and polar silanol group of silica. ...
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Analysis of reaction mixture by TLC and finding column chromatography condition

I am conducting an organic synthesis. Below is the TLC image. I used dichloromethane:methanol:triethylamine=10:1:0.15 as eluent, but I stored it during 3 days, so it may be a little bit concentrated. ...
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Is it possible to run column chromatography in reversed phase?

I'm conducting a synthesis involving aza-crown-ether. Whether aza-crown-ether acts as a base or not is unclear, but when I do TLC using dichloromethane/methanol, it is hardly drawn up with the mobile ...
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Reason for GC peak splited

I ran GCMS and one of the peak of GC appears: The peak shows some tailing, but there seems to be another peak at right-hand end of the signal. I have already checked the MS and confirmed that there ...
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Can the paper in thin layer chromatography be any type of paper?

I'm doing a chemistry chromatography lab that wants me to recreate anything I want. I decided to measure how far a pigment goes in each color for the same brand, but I'm stuck at the paper part. I ...
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Normal phase dead time marker

In reverse-phase HPLC, Thiourea, Uracil, Nitromethane or $\ce{KNO3}$ are used as dead time markers because of their polarity and therefore weak interactions with the stationary phase. However, I could ...
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Why is Retention factor for Iodide greater than Bromide?

I did a separation of Bromide and Iodide ions using paper chromatography technique for 0.3% solutions of Bromide and Iodide of sodium. I got the Retention factor equal to ~0.71 for Iodide and ~0.47 ...
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Is it possible to functionalize silica gel as Activated Carbon can be? How?

Activated carbon and Silica gel both have a high surface area. If I understand correctly it is possible to trap stuff inside activated charcoal pores to give it specific retention properties (by ...
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Comparing retention time of acetic acid, salicylic acid and acetylsalicylic acid in HPLC

I want to compare retention time of acetic acid(AA), salicylic acid(SA) and acetylsalicylic acid(ASA) in HPLC. What are the factors do I have to consider when comparing? Considering dipole moment only,...
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Can acetone be used as a GC–MS solvent for a sample containing ammonia?

Сan acetone be used as a GC–MS solvent/diluent for a sample containing ammonia? Will there be a reaction between these two species?
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Source for UV cut-off values of buffers

When I want to check a $\mathrm{p}K_\mathrm{a}$ value I look it up in CRC Handbook. Is there such source for UV cut-off values of various buffers? Usually books in chromatography contain tables with ...
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Why do I see two peaks for methyl salicylate in my GC–MS spectrum?

I ran a neat sample of methyl salicylate (CAS: 119-36-8) on the GC–MS and used $m/z~120$ as the quantifier and $m/z~92$ and $152$ as qualifiers. Below is the SIM spectrum for these ions, with two ...
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Bad repeatability of GC-FID meassurements

I am new to GC-FID and I am struggling with calibration curve. And I am strugling with the repeatability of measurements. One vial measured 5× in a row provides results which differs even 2 orders of ...
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How the plate height is assumed to be the standard deviation?

Fundamentals of Analytical Chemistry book states that the plate height can be thought of as the length of column that contains a fraction of the analyte that lies between $L$ ...
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Can I use silica gel to perform column chromatography at home?

While this question has been asked and answered, it may indicate that a column chromatography qualitatively is possible. At home, is it possible after collecting silica gels from small bags of them ...
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What are quantifier and qualifier ions in mass spectrometry data for GC-MS or LC-MS?

I've read that one should pick an ion peak that is unique to an analyte for quantification purposes and then pick at least 2 qualifier peaks and use their ratios to identify the analyte (or avoid ...
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1 answer
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Retention Time Change in Reversed Phase Chromatography (revised)

I previously learned in books that in reversed phase chromatography, retention time changes depending on the content in mobile phase, which means as the polarity of mobile phase increases, retention ...
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7 votes
3 answers
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What would a (gas) chromatogram look like if two compounds have the same retention times? [closed]

In gas chromatography, the number of peaks represents the number of compounds in an unknown sample, as retention times for each component differ. But what would happen if two components have the same ...
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How do I interpret the results of this chromatogram and does it tell me about the compounds' polarities?

I am reading this journal article which discusses using high-performance liquid chromatography (HPLC) to separate dyes from wine. Here are the relevant facts about the experimental setup: Two mobile ...
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ELSD detection principle

While documenting myself on the detectors used in HPLC, I came across one that I did not know: ELSD. I understood that the sample with the mobile phase was nebulized at the column outlet but I do not ...
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1 answer
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What are the benefits of using zero capillary voltage for discarding fractions in LC–ESI–MS/MS?

In the second episode of The Association for Mass Spectrometry & Advances in the Clinical Lab (MSACL) podcast, namely Getting going with mass spectrometry : Josh learns chromatography (aired 2021-...
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2 votes
1 answer
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How can over grinding lead to reduced API extraction in HPLC analysis?

I am looking for some advice regarding particle size distribution (PSD) & how this can be affected by grinding a tablet. During an assay by HPLC, the method requires grinding of a tablet before ...
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Right conductivity of protein sample for Protein A purification of human IgG?

A sample from a cell culture has some IgG and this is to be purified using protein A chromatography. Based on the chromatography handbook from GE-Healthcare/Cytiva, the protein sample should have &...
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4 votes
2 answers
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HPLC: Peak height calculation

I am trying to implement a small python script to detect peaks and calculate the corresponding area. The y-values are in mAU and x-values in minutes but there are some unit conversions in most HPLC ...
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