<h4>Pre-Installation Checklist</h4><ul><li>Replace oxygen, moisture and hydrocarbon traps as necessary.</li><li>Check gas cylinder pressures to ensure that an adequate supply of carrier, make-up and fuel gases are available. Carrier gases should be of the highest purity.</li><li>Ensure that the injection port is clean and free of sample residues, septum, or capillary debris.</li><li>Check and replace as necessary critical injector components such as seals, liners, and septa.</li><li>Check and replace detector seals as necessary.</li><li>Carefully inspect your column for damage or breakage.</li></ul>Note: It is critical that oxygen and water be removed from the carrier gas by the appropriate use of filters and adsorbents<h4>Injector Installation</h4>Note: GC columns do not have a specific directional flow when received from the manufacturer. Upon initial use of your new GC column, it is recommended to dedicate one specific end of the column for injector installation only. This is particularly important when dealing with active/caustic or contaminating compounds. If these compounds are routinely injected onto the column, degradation of the phase will occur—leading to higher bleed. A typical first step to remedying (removing) this bleed would be to trim 10 cm from the front (injector) end of the column and keep trimming this inlet end of the column as necessary. Trying to remedy any bleed issues by trimming the column may not work if both ends have been interchangeably installed into the inlet.<ol><li>Place a capillary nut and ferrule on the injector end of the GC column, allowing a section of column to protrude. Trim one to two centimeters from the protruding end to remove ferrule contamination that may have entered the column. Inspect the cut with a magnifier to ensure that a smooth, clean, square-cut edge has been made - recut if necessary.</li><li>Carefully hang the column in the GC oven, being cautious not to scratch or damage the polyimide coating on the capillary tubing. Rotate the column to avoid sharp bends of the capillary column and any contact of the column with oven surfaces.</li><li>Insert the column into the injector exactly the correct distance specified in the instrument manual. Tighten the ferrule nut finger-tight then 1/2 turn with a wrench. If the column can still be moved, tighten another 1/4 turn until the column is secure.</li><li>Adjust the carrier gas to obtain the flow rate listed on the test chromatogram.</li></ol>

Gas Chromatography (GC) Column Installation - Injector Installation

A buffer is an AQUEOUS solution containing a weak acid and its conjugate base (or a weak base and its conjugate acid). The pH of a buffer solution changes very little when a small amount of strong acid or base is added to it and can help to negate condition changes in an analytical method when small environmental factors vary. Buffer solutions are used to maintain the pH at a nearly constant value. They are resistant to pH change due to the presence of an equilibrium between this weak acid (HA) and its conjugate base (B).How a buffer worksWhen some strong acid (more H+) is added to an equilibrium mixture of the weak acid and its conjugate base, the equilibrium is shifted to the left, in accordance with Le Chatelier's principle. Similarly, if a strong base is added to the mixture, the hydrogen ion concentration decreases by less than the amount expected for the quantity of base added.When preparing a buffer there are a few general guidelines you should adhere to:<ul><li>Calibrate pH meter; ideally at pH values above and below target for your buffer preparation. There are calibrated buffer solutions sold for this purpose</li><li>Weigh out buffer salt to give desired buffer concentration</li><li>Add to volumetric flask containing HPLC grade water and mix thoroughly to dissolve</li><li>Using a pH probe and pH meter, measure the pH of the solution</li><li>Adjust the pH to the desired value by additions of small volumes of strong acid and/or strong base</li><li>Once the pH is correct, dilute the solution to the final desired volume and mix well</li></ul>In HPLC, buffers are typically used to control secondary interactions which are generally regarded as ‘unwanted’ in the chromatographic world. They are also used to control the ionization state of the analyte to ensure it is not present in more than one ionized state. The pKa of a compound presents the pH at which it is 50% ionized and 50% neutral. If you work at this pH you will see split peaks and variable retention times. All methods should strive to work at least two pH units away from the pKa of their target analyte.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Phenomenex_Vyznam_spravne_pripravene_mobilni_faze_822a0786d0.jpg" alt="Phenomenex: Figure 1" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Phenomenex_Vyznam_spravne_pripravene_mobilni_faze_822a0786d0.jpg 245w," sizes="100vw" width="960" height="521"><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Phenomenex_Vyznam_spravne_pripravene_mobilni_faze_2_e5b5a8b4c3.jpg" alt="Phenomenex: Figure 2" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Phenomenex_Vyznam_spravne_pripravene_mobilni_faze_2_e5b5a8b4c3.jpg 245w," sizes="100vw" width="960" height="320">When preparing isocratic mobile phases there are a few further considerations to ensure good method robustness<ul><li>Pre-Mix your mobile phase where possible, this will eliminate error from the pump</li><li>Always use the same method for preparation; for example, if you add the water first, you should always add it first. If you measure the individual components and then mix, you should always do it in this way. Do not change this process as the results can vary significantly in terms of analyte retention time.</li></ul>Below is an experiment which shows this variability when the mobile phases were mixed in the following ways<ol><li>Water first: 400 mL water in graduated cylinder, add to 1-Liter volumetric flask, dilute to mark with MeOH</li><li>MeOH first: 600 mL MeOH in graduated cylinder, add to 1-Liter volumetric flask, dilute to mark with Water</li><li>MeOH and Water measured individually: 600 mL MeOH in graduated cylinder, transfer to mobile phase reservoir; 400 mL Water in same graduated cylinder and transferred to mobile phase reservoir</li><li>Allow pump to mix A = Water with B = MeOH</li></ol><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Phenomenex_Vyznam_spravne_pripravene_mobilni_faze_3_51ea3293e9.jpg" alt="Phenomenex: Figure 3 - Kinetex™ 5µm XB-C18 150 x 4.6mm; 60:40 MeOH: H20 Flow rate 1.25mL/min Inj Vol 2µL" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Phenomenex_Vyznam_spravne_pripravene_mobilni_faze_3_51ea3293e9.jpg 213w," sizes="100vw" width="960" height="703">Buffer concentrations are also important, it is generally regarded that concentrations below 10mM do not offer the buffering capacity required to control silanol activity, or ionization state of a compound and 10mM would be considered the minimum effective concentration for reversed phase and HILIC. For other techniques such as size exclusion, significantly higher concentrations are required and it is not uncommon to see concentrations is excess of 100mM for these methods. Buffer concentration is something which must be carefully balanced to ensure solubility in all conditions can be achieved. When working under reversed phase conditions, you may need to limit the concentration of the buffer you are working with if the method contains high levels of organic (for example across a gradient) to ensure no precipitation of the buffer occurs.The mobile phase contributes as much to selectivity as your stationary phase and care with selection and preparation is as important as batch testing stationary phases when developing robust HPLC methods.

The importance of a well-prepared mobile phase

Each SPE mechanism/phase has a general set of solvent conditions under which SPE may be performed. Use the solvents/pH conditions listed below, volumes as determined in <a target="_blank" rel="noopener noreferrer" href="https://discover.phenomenex.com/0623-spe-technical-tip-part2-en?elqTrackId=E9D53E5725E641C0655E0409B9A065B7&amp;elq=00000000000000000000000000000000&amp;elqaid=4878&amp;elqat=2&amp;elqCampaignId=&amp;elqak=8AF59AC35AC6CFF8178E68A8DF72C3B8D482CF7C906A90E1BF5C55FDC1EA7D8B80B0">Method and Sorbent Volume Selection</a>.<h4>Reversed Phase SPE Method</h4><h5>Sorbent - SDB-L, C18, C8, PH, CN, X, XL</h5><ul><li>Analyte Properties: Low to moderate polarity (or nonpolar) Hydrophobic Neutralized/uncharged – Pharmaceuticals, Pesticides, Herbicides</li><li>Sample/Matrix: Aqueous, diluted with buffer – Biological fluids, Water</li><li>Conditioning Step<ul><li>Solvation – polar organic solvents – Methanol</li><li>Equilibration – aqueous, buffers – Water or buffer</li></ul></li><li>Wash Step: Aqueous buffers with 5 to 50 % polar organic solvent – Methanol:Water (5:95)</li><li>Elution Step: Polar or non-polar organic solvent(s) with or without water, buffer and/or strong acid or base – Methanol:Acetonitrile (50:50)</li></ul><h4>Normal Phase SPE Method</h4><h5>Sorbent - Silica, Florisil®, NH2, CN</h5><ul><li>Analyte Properties: Moderate to high polarity compounds (neutralized/uncharged) – Pesticides</li><li>Sample/Matrix: Non-polar organic solvents or moderately polar organic solvents – Hexane, chloroform, petroleum ether, toluene or methylene chloride</li><li>Conditioning Step<ul><li>Solvation – polar organic solvents (optional) – Methanol (optional)</li><li>Equilibration – sample/matrix solvent – Hexane or chloroform</li></ul></li><li>Wash Step: Non-polar organic solvents with a low concentration (1 to 5 %) of moderate to low polarity organic solvents – Hexane with 1 % THF, ethyl acetate, acetone, acetonitrile or IPA</li><li>Elution Step: Non-polar organic solvents containing higher concentrations (5 to 50 %) of moderate to high polarity organic solvents – Hexane with 10 % THF, ethyl acetate, acetone, acetonitrile or IPA</li></ul><h4>Strong Ion-Exchange SPE Method</h4><h5>Sorbet:&nbsp;Anion exchange: Screen-A, SAX, X-A, X-AW, XL-A, XL-AW Cation exchange: Screen-C, SCX, X-C, X-CW, XL-C, XL-CW</h5><ul><li>Analyte Properties: Ionized/charged compounds – Anion exchange: Acidic analytes Cation exchange: Basic drugs</li><li>Sample/Matrix: Aqueous; Low ionic strength buffers (&gt;30 mM), pH adjusted – Biological fluids plus buffer</li><li>Conditioning Step<ul><li>Conditioning – polar organic solvents – Methanol</li><li>Equilibration – low ionic strength buffers, pH adjusted – 25 mM Tris-OAc, pH 7</li></ul></li><li>Wash Step: Aqueous buffers of low salt concentrations with or without organic solvent<ul><li>Anion exchange: Buffer pH 7: Methanol (50:50)</li><li>Cation exchange:<ul><li>Buffer pH 6</li><li>1 M acetic acid</li><li>3. Methanol</li></ul></li></ul></li><li>Elution Step:<ul><li>Neutralize the charge on the weak anion or cation</li><li>Increase the ionic strength and counter ion concentration</li><li>Add a strong counter ion displacer<ul><li>Anion exchange: Hexane:ethyl acetate (75:25) + 1 % glacial acetic acid</li><li>Cation exchange: Methanol + 5 % NH₃</li></ul></li></ul></li></ul><blockquote>NOTE: These are general starting conditions. Please <a target="_blank" rel="noopener noreferrer" href="https://discover.phenomenex.com/customersupportEU?elqTrackId=9FD41A83311DD308567C001F2CC9485A&amp;elq=00000000000000000000000000000000&amp;elqaid=4878&amp;elqat=2&amp;elqCampaignId=&amp;elqak=8AF530A7A08EE95226A9F3C49828A74B76F7CF7C906A90E1BF5C55FDC1EA7D8B80B0">contact </a>your Phenomenex technical consultant for more assistance.</blockquote>

Generic SPE Method by Retention Mechanism

Article

Product

Events

Webinars

Application

Academy

Video

Poster

Scientists

Plastics

Environment

Food

Water

Health

Presentation

Scientific article

Interview

Toxicology

Popularization

Science and research

Software

Covid-19

Career

Laboratories

Various

PFAS 

Zaměření

Type of News

Focus

Phenomenex is a global technology leader committed to developing novel analytical chemistry solutions that solve the separation and purification challenges of researchers in industrial, government and academic laboratories.

Generic SPE Method by Retention Mechanism

The importance of a well-prepared mobile phase

Gas Chromatography (GC) Column Installation - Injector Installation

Upcoming events

Bruker at SOFT-TIAFT 2026

Thermo Fisher Scientific at SOFT/TIAFT 2026

Thermo Fisher Scientific Environmental & Food Safety Workshop Series 2026

Webinars

How to Perform Routine Quantification by FTIR: Tips & Tricks Workshop

Learn How FTIR Can Help You with Contamination and Poor Product Quality

Expanding the Evidence: New Data for More Reliable PFAS Analysis