Chromatography

Syringes

Syringe Cleaning and Maintenance

Chromatography syringes are the finest quality precision fluid measuring devices available. With proper care and handling, syringes will provide unsurpassed performance in precision fluid measuring year after year. The life of your syringe is directly related to its cleanliness!

Some solvents, such as halogenated hydrocarbons, may attack and deteriorate the highly resistive adhesives (cements) used to affix needles and other terminations to Hamilton syringes, which may result in frozen plungers and plugged needles.

Cleaning Syringe Barrels

To clean Hamilton syringes, it is best to use solvents known to be effective in solvating the sample and preferably are non-alkaline, non-phosphate and non-detergent based. A biodegradable, non-phosphate, organic Cleaning Concentrate is available from Hamilton (ordering number 18311).

Rinse the syringe thoroughly after use with deionized water, acetone, or another solvent compatible with the sample. Allow the syringe to air dry. Avoid prolonged immersion of the syringe while cleaning.

MICROLITER™ Syringes (Series 600, 700, 800 and 900)

• Rinse the syringe thoroughly with a solvent known to be effective in solvating the sample. Residual dissolved solids may result in frozen plungers and plugged needles.
• To clean the plunger, remove it from the syringe barrel and gently wipe with a lint-free tissue. Reinsert the plunger into the barrel and pump deionized water, acetone or another solvent compatible with the sample through the needle and syringe. Allow syringe to air dry. When working with dissolved solids, storing the plunger outside of the syringe will reduce the possibility of frozen plunger.

GASTIGHT® Syringes (Series 1000, 1700 and 1800)

• Rinse the syringe thoroughly with a solvent known to be effective in solvating the sample. Residual dissolve d solids may result in frozen plungers and plugged needles.
• To clean the plunger, remove it from the syringe barrel and gently wipe with a lint-free tissue. Insert the plunger into the barrel and pump deionized water, acetone or another solvent compatible with the sample through the needle and syringe. Allow syringe to air dry. When working with dissolved solids, storing the plunger outside of the syringe will reduce the possibility of frozen plunger.

Syringe Storage

We recommend to store syringes in the original packaging. This with help to protect the syringe, and allows for easy identification. Remove product description label from the end of the box, and placing it to the outside packaging. This will make re-ordering the same syringe quick and easy.

Thermal desorption

In this section you will finde informations important in the area of thermal desorption. It is quite difficult analytical technique and these informations can help you with your work with it. If you do not find required information, do not hesitate to contact our specialists.

Sorbent parameters

Material emission monitoring

Storage and transportation of sorbent tubes

ChromShell columns and solvents

There are several critical characteristics that must be taken into consideration when selecting the appropriate organic solvent to use in the mobile phase with ChromShell^® columns. Viscosity is one of the most important as high viscosity solvents may produce backpressures that are too high for the HPLC system used. Other important solvent characteristics include UV cutoff, cost and polarity index; where a solvent with a high UV cutoff will result in poor sensitivity with UV/Vis detection and use of high cost solvents will result in a poor laboratory that can't afford to buy new columns. Solvents with low polarity indices generally result in faster elution of organic compounds and are commonly used for column cleaning.

Acetonitrile

is arguably the best organic solvent as it results in the lowest system backpressure in water mixtures and also has a very low UV cutoff for better UV/Vis detection sensitivity. Although acetonitrile production is starting to increase with the turnaround of the economy, thus dropping cost, this still remains a major drawback of acetonitrile usage.

Methanol

is another popular organic solvent as it is comparable in elution strength to acetonitrile, has a relatively low UV absorbance, and is significantly less expensive than acetonitrile. The major drawback of methanol, especially when used with small particle size HPLC columns, is that its use can result in backpressures that exceed many HPLC system limits.

Acetone

is less commonly used as it has high UV absorbance, but can be used successfully if analytes absorb at higher UV wavelengths or if other detector types such as MS are used as it has similar elution properties to acetonitrile but is significantly less expensive.

Ethanol

is generally not recommended as it results in very high backpressures in water mixtures.

Iso-, n-propanol

have relatively strong elution strength and are most commonly used in column cleaning at low flow rates as they also results in high backpressures.

Tetrahydrofuran

has similar elution strength to n-propanol but is less commonly used as it is much more expensive.

Glassware deactivation

Glassware deactivation with DMDCS

Dimethyldichlorsilane (DMDCS) reacts with active hydroxyl groups present on glass surface producing a deactivated layer. This ensures inert glassware used for sensitive compounds.

Procedure

During the deactivation process, the reaction releases hydrogen chloride (HCl). So we strongly recommend to do this procedure in a fume hood.

• Use 5% DMDCS solution in toluene. You can prepare this solution by diluting 20 ml of DMDCS with 400 ml toluene. Store the solution in an amber glass at room temperature.
• Soak glassware in 5% DMDCS solution for 15 to 30 minutes.
• Rinse glassware twice with toluene.
• Soak glassware in methanol for 15 minutes.
• Rinse glassware with methanol.
• Dry glassware with high purity nitrogen (moisture and hydrocarbon free).

Setting linear velocity

The linear velocity is an important parameter in chromatography. It has an influence on chromatography resolution and therefore setting the dead volume is basic part of method development.

Linear velocity measurement in GC

To set a dead time, inject 2 µl of a non-retained gaseous substance, which is compatible with the detector. Take a gas-tight syringe and draw the headspace over neat compound. Accurately mark the injection starting time and peak elution time.

Note: Some compounds may be slightly retained on thick-film phase, however, they will be reproducible for similar column types.

Handling chiral columns

Cellulose/Amylose chiral column use and care

Shipping Solvent

n-Hexane/2-propanol (9:1, v/v)

Test Certificate

Each column is individually tested before shipment. A test certificate showing the separation parameters for trans-stilbene oxide is enclosed with each column.

Mobile Phase Compatibility

Chiral columns can be used with normal phase (alkane/alcohol), reversed phase (aqueous methanol, aqueous acetonitrile or appropriate buffer/methanol or buffer/acetonitrile mixtures), as well as with pure polar organicsolvents (low molecular weight alcohols, acetonitrile or their mixtures).

Solvent Switching

An appropriate column washing procedure must be applied when changing from one mobile phase to another. The miscibility of the different mobile phase components must be carefully considered for this wash. To safely transfer a column from hexane to methanol (or acetonitrile) or from methanol (or acetonitrile) to hexane, use 100 % 2-propanol as transition solvent at a flow rate of 0.2-0.5 mL/min. Ten column volumes of 2-propanol (i.e. 25 mL for a 250 x 4.6 mm i.d. column or 15 mL for a 150 x 4.6 mm i.d. column) are sufficient for completely removing the old mobile phase. To safely transfer a column from normal phase to reversed phase conditions flush the column with 100 % 2-propanol at 0.2-0.5 mL/min for minimum ten column volumes. In addition, when the buffer salt additive of the RP mobile phase is insoluble in 2-propanol, flush the column briefly with water before switching to a buffered mobile phase. We recommend the use of dedicated Lux columns to reversed phase operation hence avoiding the need of converting columns used in normal phase elution mode to reversed phase or vice versa.

Use of Mobile Phase Modifiers

For basic samples or acidic chiral compounds, it may be necessary to use an appropriate mobile phase modifier in order to achieve chiral resolution and to insure proper peak shapes. Diethylamine, ethanolamine and butyl amine in the concentration range 0.1-0.5 % can be used with basic analytes, while trifluoroacetic or acetic acid (0.1-0.5 %; typically 0.1-0.2 %) with acidic analytes. Mixtures of basic and acidic mobile phase additives are acceptable (e.g. diethyl amine acetate or trifluoroacetate). Lux columns will deliver consistent results when operated with mobile phases containing additives at the concentration levels specified above. However, limited decrease in column efficiency may occur when a column is used in combination with these additives. Therefore, we advise to dedicate columns to mobile phases containing basic additives. Mobile Phase Restrictions Lux chiral stationary phases are prepared by coating silica with various polysaccharide derivatives. Therefore, any solvent dissolving the polysaccharide derivative (such as tetrahydrofurane, acetone, chlorinated hydrocarbons, ethylacetate, dimethylsulfoxide, dimethylformamide, N-methylformamide, etc.) must be avoided even in trace amounts (e.g. even as sample solvent).

Operating Backpressure

The mobile phase flow rate should be set such that the column backpressure stays below 300 bar (4300 psi). This maximum backpressure should not be exceeded for long periods of time.

Operating Temperatures

With standard mobile phases (such as alkane/alcohol) the column can be used in the temperature range 0-50 °C.

Column Storage

Column storage for a longer period of time is recommended in n-hexane/2-propanol (9:1, v/v). Columns used in reversed phase conditions should be first flushed with water (whenever a buffer salt was used as RP mobile phase additive) and then with methanol (or with methanol only when no salt was used). The column can be stored in methanol.

Extending Lifetime and Reconditioning

We recommend the use of guard cartridges to extend the lifetime of your column, especially with samples extracted from complex matrixes. Ideally, samples must be completely dissolved in the mobile phase or filtered through a syringe filter of approximately 0.45 μm porosity.

Hints and tips

This site includes technical information, hints and tips for your decisions, and recommendation for selecting right chromatography accessories.

• Agilent GC inlet seal improvment
• Dynamic headspace
• GC column conditioning procedure
• GC column installation
• GC liners
• GC septa selection
• Glassware deactivation procedure
• How to improve GC capillary column connections
• How to increase signal/noise ratio in GC systems
• How to set optimum linear velocity
• How to handle chiral HPLC columns
• LC separation performance improvement
• LC column selection
• Syringe care
• Syringe tip selection
• Thermal desorption

Dynamic Headspace

Dynamic Headspace (DHS) can be used for a wide range of environmental applications. Pollutants can be detected in drinking water, river water, and waste water. These matrixes include chemicals of different polarity, volatility, such as chlorinated hudrocarbons, aromatics, oxygenates, etc. Dynamic Headspace also can be used for further applications:

• characterization of spices, herbs, foods, soaps
• residual monomer and other VOC in polymers
• residual solvents in food packaging
• "green label" product testing
• OVIs in pharmaceutical products
• trace impurities in active pharmaceuticals ingredients
• metabolites in biological fluids (aromatic hydrocarbons in urine, benzene in blood)

Capillary GC column installation

Brief procedure for column installation

• Cool all heated zones of GC
• Check gas purifiers and if they are spent, replace them.
• Clean injector and detector
• Replace injector/detector liners with new ones.
• Replace critical injector and detector seals.
• Replace septum in the injector.
• Set make-up and detector gas flow rates.
• Carefully inspect the column for damage or breakage.
• Install a nut and ferrule on each end of the column.
• Cut 10 centimeters from each end of the column. Use a sapphire scribe or ceramic scoring wafer to cut fused silica capillary columns. Use the serrated edge of a ceramic scoring wafer or the edge of a sharp file to cut metal capillary columns. See our catalogue for capillary cuting tools.
• Mount the capillary column in the oven using a bracket that protects the column from becoming scratched or abraded.
• Insert column the appropriate distance into the inlet as indicated in the instrument manual.
• Install the column so that the capillary does not touch the oven walls.
• Set the approximate column flow rate by adjusting the head pressure to the value listed on the test chromatogram included with the column.
• Set split vent, septa purge, and any other applicable inlet gases according to the instrument specifications.
• Confirm the flow by immersing the column outlet in a vial of solvent (acetone or isopropyl alcohol).
• Insert column the appropriate distance into the detector as indicated in the instrument manual.
• Check for inlet and outlet leaks using a thermal conductivity leak detector. Do not use soaps or liquid-based leak detectors or the column may be damaged.
• Set injector and detector temperatures. Turn the detector on when the temperatures have equilibrated. Caution - do not exceed the phase's maximum operating temperature!
• To set the proper dead time (linear velocity), inject methane or a non-retained substance compatible with the detector being used.
• Verify system integrity by checking the dead volume peak. It should not tail.
• Condition the column at its maximum operating temperature to stabilize the baseline. (See the test chromatogram included with the column for the maximum temperature.)
• Set oven to appropriate temperature and inject methane or an appropriate unretained substance, again to set the proper linear velocity.
• Inject a duplicate of the original test mixture or your specific test mixture to confirm proper installation, system, and column performance.
• Calibrate the instrument and inject samples.

Note: If the column is new, you have to run conditioning procedure prior setting the proper dead time.

Unit Conversions

HPLC capillaries volume chart