Questions tagged [chromatography]

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

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55 views

Organic solvent for cleaning autosampler syringe

I work in a laboratory where gas chromatography is used extensively. We handle phthalates. What are the possible wash solvent for autosampler syringe? I've been using acetone.
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26 views

Column Dispersive Mass Balance analytical / numerical solution

Is there any analytical solution for the most common expression of column mass balance: $\dfrac{\partial C}{\partial t} + \left(\dfrac{1+\varepsilon}{\varepsilon}\right)\dfrac{\partial q} {\partial t} ...
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555 views

Typesetting of the retardation factor (Rf)

What should be the typesetting for the symbol of the retardation factor as used in thin-layer chromatography? These are the possibilities: Rf - only f in italics Rf - both R and f in italics Rf - ...
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61 views

When to use chromatographic separation vs vacuum-assisted rectification distillation?

I'm trying to understand a bit more about industrial scale separation processes. One particular element I'm interested in is the question as to when chromatography (HPLC or Simulated Moving Bed) may ...
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1answer
165 views

Stronger ESI signal for deuterated substances

I observe a constant phenomenon in my ESI measurements, which indicates that deuterated substances give a stronger signal than the same substance in non-deuterated form. Unfortunately I can't make any ...
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2answers
28k views

Normal phase vs reverse phase HPLC

What is the difference between normal phase and reverse phase high-performance liquid chromatography?
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67 views

Is there any database, or reference, for retardation factors (Rf)? [closed]

I'll like to compare the Rf of myrcene, obtained in Thin Layer Chromathography, for different species. Toluene:Ethyl Acetate (97:3) would be fine, but is not a necessity.
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1answer
52 views

Theoretical plates in Plate theory of chromatography

In "Chromatography theory" by Scott and Cazes, The plate volume is given as ($v_m+Kv_s$) where $v_m$ and $v_s$ are the volume of stationary and mobile phase in the column and $K$ is the ...
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0answers
35 views

How do I analyze 17 different gases from a sample?

I am designing a test setup in which I heat a battery until it releases its gases, and I need to analyze the composition of the fumes. Below is the list of gases that are believed to be present in the ...
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2answers
1k views

Plotting a calibration curve for Gas-Liquid Chromatography

Question: The given graph of abundance of different samples of butanol with respect to time is- Textbook solution: But why aren't the points like this: $(20,5);(40,10);(60,15);(a,7)$ after which, ...
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1answer
28 views

Elution and void volumes in a gel chromatography experiment

I was using gel-filtration for separation Hb from xylene cyanol FF with Blue Dextran to determine the void volume. Sephadex G-25 is stationary phase, and mobile phase was buffer $(\pu{0.020 mol dm-3}$ ...
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1answer
2k views

Analytical Chemistry predicting the order of elution of compounds in a reverse phase HPLC

Predict the order of elution for a mixture containing caffeine and benzoic acid, using a reverse phase HPLC. Explain your answer. My try: Since benzoic acid is more acidic that caffeine, which means ...
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2answers
106 views

Can separated and purified substances be obtained from a chromatogram?

I am studying chemistry, and I am learning about chromatography. But I am confused about how chromatography is used to ‘separate and purify’ substances. I have learnt about chromatography paper, ...
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20 views

Disadvantages of SIM vs TIC in GC-MS [closed]

In GC-MS what are the relative advantages/disadvantages of using Selected-Ion Monitoring (SIM) vs using Total Ion Chromatograms (TIC)? I know that SIM offers a better S/N ratio, however if the S/N is ...
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1answer
108 views

Software platforms with integration of multi vendor mass spectrometer device data?

I know about ACD/Spectrus Processor which is a software platform to integrate analytical chemistry data from multiple mass spectrometer device vendors (e.g. Agilent Technologies, Bruker, Waters) to ...
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58 views

Purification of primary amines using Schiff base immobilization

I want to purify all primary amine containing compounds in a complex mixture (cell extracted metabolites). I am not aware of any published methods to do so, so I am thinking to make my own using ...
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1answer
66 views

How do I import CSVs to OpenChrom?

My end objective is to compare GC outputs of samples in two groups using PCA. I have bene given the data in Excel format but have saved to CSV. Whilst I can locate CSV files through OpenChrom, they ...
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1answer
50 views

Why can DAP be used to elute a protein in affinity chromatography?

A little background: as part of my bioinformatics degree I have to take Protein Chemistry course, but I miss chemistry basics (have CS background), so that's why I'm asking the following question. We ...
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1answer
81 views

Is there a list of must-have columns for LC-MS/MS used in biochemistry/clinical lab setting?

Considering a wide variety of chromatographic columns used for (U)HPLC based on phase type (normal or reverse); column dimensions (geometry, length, inner diameter); particle material $(\ce{SiO2},$ $\...
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1answer
48 views

Why can zwitterions be difficult detect by HPLC-MS?

I understand that to separate molecules by HPLC (or SFC) you commonly adjust the pH of the mobile phase to get preferable charges (i.e. negative charges) on the molecules - so that they interact with ...
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1answer
36 views

What order would these compounds elute in a 3:1 hexanes:ethyl acetate TLC run? [closed]

I would figure the order would be (in order of smallest Rf to largest) W, Z, Y, X. Is the carboxylic acid more polar than the acetic anhydride?
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1answer
93 views

Conditions for choosing quantifying and qualifying ion in LC-MS/MS

In my PhD-work, we are trying to develop LC-MS/MS methods for targeted analysis. Usually, I determine my quantifying ion/fragment based on a few different criteria: If there is a 13C internal ...
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3answers
8k views

Suitable solvents for column chromatography

Why are 100% hexane or 100% ethyl acetate never used as eluents in column chromatography?
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2answers
153 views

What is the principle of RP-HPLC? [duplicate]

Is the following statement the correct way to explain principle of RP-HPLC? It works on the principle of hydrophobic interactions hence the more nonpolar the material is, the longer it will be ...
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1answer
189 views

How to pool fractions after chromatography?

I used chromatography to purify my protein and now I have fractions with different concentrations ranging from 1-16 mg/ml. Should I pool all these fractions or just the ones with high concentration (...
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1answer
81 views

How to recover C18 column after prolonged exposure to phosphate buffer?

A colleague of mine who occasionally does HPLC in a biochemistry lab discovered that a C18 column was not properly flushed after the previous experiment. It appears that the column was stored for ...
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1answer
60 views

Can we use HPLC to purify an organic reaction product?

I know HPLC is a form of column chromatography that uses high pressure to pump a sample mixture/eluent through a column. However, I'm not sure if this technique is being used to purify and isolate ...
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1answer
84 views

Protein purification with Cobalt

Cobalt exhibits a more specific interaction with histidine tags, resulting in less nonspecific interaction than nickel. For this reason, cobalt is the preferred divalent cation for purifying His-...
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1answer
90 views

Why n-alkanes can be found in organic solvent extract of rubber?

Sorry for the confusing title, I didn't know how to describe this better. So, if we douse rubber, say, a bicycle tire or a certain type of rubber glove, with organic solvent (n-heptane), we can find n-...
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1answer
802 views

Different elution profiles of carvacrol and thymol

I have identified carvacrol and thymol via thin-layer-chromatography and via gas chromatography On a normal silica plate the thymol has a higher Rf-value then carvacrol. The explanation for this would ...
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2answers
509 views

What dictates cathode vs anode nomenclature use?

what was the justification for the use the cathode and anode terminology in IEF? There is no redox taking place in IEF, just proteins interacting with an electric field
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50 views

Why mustn't the sample be dipped in the mobile phase in paper chromatography?

As the title says it, I am curious about why the sample (ie, an extract of plant's chlorphyl transferred onto paper) must not be dipped into the mobile phase (ie, the solvent which goes up the paper)? ...
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3answers
547 views

In High Performance Liquid Chromatography, why are ratios of solvents used?

How do they allow for better separation? For example, why isn't 100% hexane used?
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125 views

Choosing LC-MS modifier - why only acids?

During method selection our chromatographists have options like NH4, TFA, Formic Acid. What's the point of choosing only between acid? Why not use the strongest that guarantees ionization? The sample ...
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2answers
60 views

Can I normalize my data in order to compare it?

I have three chromatograms of the same eluted protein but at different concentrations, since it got diluted during purification process) and hence peak maximums at very different absorbances. The ...
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1answer
2k views

Single Point vs. 5 Point Calibration in HPLC

Single-Point calibrations aren't that useful for quantitative analysis and there is more room for error in result readings as compared to a 5-Point calibration. Is this why a 5-Point calibration is ...
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I am having difficulties with analysing steroids in GC/MS [closed]

Is there anyone here that has already analysed estrone and 17beta-estradiol on GC/MS (HP5-MS column and MSTFA as derivatizing agent)? Is it necessary to make a mixture of other reagents with MSTFA or ...
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1answer
40 views

How appropriate/adaptable is HPLC for in situ sample analysis on Titan?

I have a limited background in chemistry (nothing beyond general Chem 2), so I am asking for help with a group project. We have two chemists on our team, but communication is limited at the time and I ...
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33 views

Protein purification pressure limits confusion, about the X-MPa limit for FPLC?

I have always been a bit confused about column pressure limits. I have to elaborate a bit on the problem to get to my question: In protein purification we use the äkta purification machines (e.g. ...
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Do I have to change my sample buffer before IEX chromatography?

I will use IEX chromatography to purify a protein and will then use a equilibration buffer Tris-HCl, pH 8 to set the start conditions for my column and optimize the binding of my protein. Then inject ...
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Change buffer in dialysis?

I have a protein that I purified in PBS buffer, pH 7. I will do dialysis to remove salt and will then further purify the protein with ion exchange chromatography. I will need to use another buffer (...
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1answer
94 views

Column length for baseline separation

I have a GC-chromatogram with 4 retention times and peak widths and a column length. How can I calculate the column length that I need in order to get baseline separation for all components in the ...
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30 views

Acid and base separation on an anion exchange column [closed]

I'm new with this concept and so I would need some advice. I want to separate an acid and a base in a water solution. The acid has $\mathrm{p}K_\mathrm{a}(\ce{HA}) = 5$ and the corresponding acid of ...
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3answers
94 views

Applying vacuum at the end of a flash chromatography column

I know that flash chromatography uses a positive or pushing pressure to achieve the goal which is separation. Would the same goal be achieved if you had a vacuum on the other end instead of a ...
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1answer
17k views

How to determine the order of elution in silica gel column chromatography?

Benzofuranoids ustusoranes A (1), B (2), and E (3) were isolated from Aspergillus ustus 094102. If these compounds were separated by column chromatography on silica gel, what would be the elution ...
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1answer
72 views

Physical meaning of tm

The definition of dead time ($t_\mathrm m$) is mentioned as the time taken for elution of an unretained species. It is also mentioned that the retention time ($t_\mathrm r$) is sum of the time analyte ...
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1answer
230 views

In HPLC, how does column diameter affect the retention time?

I am using a smaller diameter column than my previous one keeping all other factors such as stationary phase, solvent, solvent gradient constant. (Obviously, flow rate will be less in smaller column). ...
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2answers
3k views

Retention time and dead time

Why does ts start just after tm ? Does the analyte starts spending time in the stationary phase only after the mobile phase has passed and reached the detector ? Wouldn't there be an overlap between ...
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Concept of Adjusted Retention Time in Chromatography

Why is the relation between the adjusted retention time, retention time and dead time as follows : tR' = tR - tM ? To find the time that the sample spends in stationary phase (adjusted retention ...
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226 views

Gas chromatography flame ionization detector (FID) - why hydrogen gas?

Why is $\ce{H2}$ used in an FID, apart from the fact its combustion does not contaminate the flame? In other words, is the temperature of the flame important or critical? Is the stability of the ...