l 2hhWaWbｔRW0.5 htR: Retention time(min)W0.5 h: Peak width at 50 % of the peak height(min)H : Peak heighttRWbWaWa: Front half of the distance where the tangent line W0.5 hWb: Back half of the distance as above (min) General Precautions for liCoumnHandng0.5 h0.1 hN = 5.54×Fas=to a Gaussian peak intersects the baseline at 10 % of the peak height (min) To achieve the best column performance, please follow the below instructions.solution with the eluent to be used.• If desired new eluent has low miscibility/solubility to the eluent of previous analysis, first use the eluent that is miscible/soluble to both eluents, and then replace it with the desired eluent. *If the eluent left in the HPLC system is not compatible with the column to be used, it may damage the column. *A drastic change in the eluent compositions may remove substances adsorbed on the HPLC system and they may enter and deteriorate the column.position the guard column in front (before the inlet) of the analytical column.• Make sure to insert the tubing all the way to the end fitting and secure it with the male nut. It is important that there is no extra space between the tubing and the column side of the end fitting. Presence of an extra space will let the sample to spread out and may result in wide peaks.• Set the initial flow rate at less than half of the recommended flow rate and start the system. If the column is to be heated during the analysis, keep the low flow rate until the column temperature reaches to the set temperature, and then gradually increase the flow rate to the desired temperature. *Verify that there is no solvent leak. The solvent leak may cause electronic leakage, rust, and/or chemical injury. *Make sure not to let air bubbles enter the column while installing the column. The air bubbles may damage the column. * When restarting the system after column installation or after holding the eluent flow, start the system at less than half of the recommended flow rate. A rapid increase in pressure can damage the column. * If the column was heated during the analysis, lower the flow rate to less than half of the recommended flow rate at the end of analysis. Then, turn off the column oven to let the column temperature returns to room temperature before stopping the pump. This is to prevent creating an empty space in the column, which deteriorates the column. Since if the pump was stopped while the eluent inside the column is still hot, the eluent volume decreases and creates an empty space when the eluent temperature decreases. *It is recommended to set the pump limiter to avoid exceeding the maximum pressure. 5 times of the column volume.• Check miscibility/solubility of the desired new solvent and the solvent currently filled in the column.• When replacing with a solvent having low miscibility/solubility to the current solvent, first use a solvent that is miscible/soluble to both (new and current) eluents, and then replace it with the new solvent.• When using a gradient method, changes in the eluent compositions may increase the column backpressure. Adjust the flow rate and column temperature so that the column backpressure remains below the usable maximum pressure throughout the analysis.the column at a location with stable temperature (a cool and dark space is recommended). *Never allow inside the column to dry. It can damage the column. • Theoretical Plate Number (N) HPLC System Preparation• Wash entire HPLC system prior to column installation, including all flow-lines and sample loop by switching the valve, and then replace the washing Column Installation• Connect the column to HPLC system by following the “flow direction arrow” ( Solvent Exchange• To replace the solvent, start the system at less than half of the recommended flow rate. Recommended solvent volume to introduce at each step is 3 to Column Storage• Remove the column from HPLC system after replacing the in-column solvent with the initial shipping solvent. Securely tighten the end caps and store Column Inspection• Inspection method is described in the Certificate of Analysis (CoA).• Theoretical Plate Number (N) and Asymmetry Factor (Fas) were calculated using the below equations. Additional Warnings• Do not remove end fittings.• Do not make a strong impact on the column. Do not drop or hit the column on a hard surface.• Please follow a proper waste disposal method specified by your local regulations. ) indicated on the column adhesive label. If guard column is used, *Read the operation manual before using the column.Injection• Asymmetry Factor (Fas)Injection* Plate count and Fas values change significantly depend on samples and/or analysis conditions being used. To check the initial column condition, please make sure to use the same sample and the analysis condition mentioned in the CoA.69General Precautions for Column Handling
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