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Agilent 1260 Infinity Quaternary LC VL Manual and Quick Reference

3

Optimization of the Agilent 1260 Infinity Quaternary LC VL

Optimizing Detection with DAD

Figure 19

Optimization of Wavelength Setting

A wide bandwidth has the advantage of reducing noise by averaging over a
wavelength range — compared to a 4 nm bandwidth, the baseline noise is
reduced by a factor of approximately 2.5, whereas the signal is about 75 % of a
4 nm wide band. The signal-to-noise ratio for a 30 nm bandwidth is twice that
for a 4 nm bandwidth in our example.

Figure 20

Influence of Bandwidth on Signal and Noise

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Содержание 1260 Infinity Quaternary LC VL

Страница 1: ...Agilent Technologies Agilent 1260 Infinity Quaternary LC VL System Manual and Quick Reference...

Страница 2: ...is document are furnished under a license and may be used or copied only in accor dance with the terms of such license Restricted Rights Legend If software is for use in the performance of a U S Gover...

Страница 3: ...cifications for the LC system 3 Optimization of the Agilent 1260 Infinity Quaternary LC VL This chapter considers how to apply the theory and use the features of the LC system to develop optimized sep...

Страница 4: ...artment 24 Detector 25 2 Specifications 37 Physical Specifications 38 Performance Specifications 40 3 Optimization of the Agilent 1260 Infinity Quaternary LC VL 49 Optimizing the Pump 50 Optimizing th...

Страница 5: ...pendix 99 Safety 100 The Waste Electrical and Electronic Equipment Directive 103 Lithium Batteries Information 104 Radio Interference 105 Sound Emission 106 Solvent Information 107 UV Radiation 108 De...

Страница 6: ...6 Agilent 1260 Infinity Quaternary LC VL Manual and Quick Reference Contents...

Страница 7: ...ary LC VL 10 System Components 11 Optimizing the Stack Configuration 12 One Stack Configuration 12 Two Stack Configuration 15 Quaternary pump 17 Hydraulic Path 18 Autosampler 19 Sequences 21 Thermosta...

Страница 8: ...ties The Agilent 1260 Infinity Quaternary LC VL is ideally suited for multi method high throughput workflows It offers Convenient access to four solvents for isocratic or gradient analysis for rapid m...

Страница 9: ...usly tracks long term instrument usage and user defined limits with feedback message when limit is exceeded Upgradeability and expandability with the complete range of Agilent 1200 Infinity Series HPL...

Страница 10: ...noise and very low acoustic noise for better results and better working environment Degasser and automatic purge valve integrated into pump module Variable volume autosampler with reduced delay volume...

Страница 11: ...d for the Agilent 1200 Infinity Series system for increased analysis speed with sensitivity resolution and precision Thermostatted column compartment This stackable temperature controlled column compa...

Страница 12: ...m in following configurations These configurations optimize the system flow path ensuring minimum delay volume One Stack Configuration Ensure optimum performance by installing the modules of the Agile...

Страница 13: ...LC VL Manual and Quick Reference 13 Introduction 1 Optimizing the Stack Configuration Figure 1 Recommended Stack Configuration Front View HdakZci XVW cZi Ejbe 6jidhVbeaZg 9ZiZXidg I ZgbdhiViiZY Xdajbc...

Страница 14: ...Introduction Optimizing the Stack Configuration Figure 2 Recommended Stack Configuration Rear View A6C id Xdcigda hd ilVgZ 6cVad YZiZXidg h cVa dg djiejih eZg YZiZXidg adXVi dc YZeZcYh dc YZiZXidg 68...

Страница 15: ...e system it is recommended to form two stacks Some users prefer the lower height of this arrangement even without the autosampler thermostat A slightly longer capillary is required between the pump an...

Страница 16: ...al and Quick Reference 1 Introduction Optimizing the Stack Configuration Figure 4 Two stack configuration rear view 68 EdlZg 86C 7jh XVWaZ GZbdiZ XVWaZ I Zgbd XVWaZ dei dcVa 86C 7jh XVWaZ id chiVci E...

Страница 17: ...ssembly which can generate pressure up to 400 bar Degassing of the solvents is done in a built in vacuum degasser Solvent compositions are generated on the low pressure side by a high speed proportion...

Страница 18: ...LC VL Manual and Quick Reference 1 Introduction Quaternary pump Hydraulic Path Figure 5 Hydraulic Path of the Quaternary Pump Id lVhiZ gdb hdakZci WdiiaZh KVXjjb X VbWZg 9Z VhhZg DjiaZi kVakZ 9VbeZg I...

Страница 19: ...ns and automatic delay volume reduction ADVR contribute to faster cycle times and even faster gradient delivery to the column The system draws exactly the set volume of sample solution without waste a...

Страница 20: ...orrect operation The metering device is always flushed after injection to ensure minimum carry over The module uses an analytical head providing injection volumes from 0 1 to 100 L for pressures up to...

Страница 21: ...ection valve returns to the mainpass position at the end of the sampling sequence The sampling sequence occurs in the following order 1 The injection valve switches to the bypass position 2 The plunge...

Страница 22: ...this position the mobile phase flows through the autosamplers metering device sample loop and needle ensuring all parts in contact with sample are flushed during the run thus minimizing carry over Fi...

Страница 23: ...sample into the sample loop Figure 9 on page 23 Figure 9 Drawing the Sample When the metering unit has drawn the required volume of sample into the sample loop the needle is raised and the vial is re...

Страница 24: ...ng from 10 degrees below ambient up to 80 C with high heating and cooling speeds for maximum application flexibility and stability Holds up to three 30 cm columns and optimized design gives minimum de...

Страница 25: ...ted due to less baseline noise drift refractive index and thermal effects especially under ultra fast gradient conditions RFID tracking technology is used for the UV lamp and the Max Light cartridge f...

Страница 26: ...UV wavelength range Its light is focused by a lamp mirror 2 onto the entrance of the Max light cartridge flow cell 3 with optofluidic waveguides The light leaves the Max light cartridge flow cell at...

Страница 27: ...Introduction 1 Detector Lamp The light source for the UV wavelength range is a long life UV lamp with RFID tag As a result of plasma discharge in low pressure deuterium gas the lamp emits light over t...

Страница 28: ...Cartridge Test Cell is available The optical principle of the Max Light Cartridge cell is based on opto fluidic waveguides Nearly 100 light transmission is achieved by utilizing total internal reflec...

Страница 29: ...d by using a concave holographic grating The grating separates the light beam into all its component wavelengths and reflects the light onto the photodiode array The diode array is a series of 1024 in...

Страница 30: ...depending on the application needs other types may be introduced later easy front access to lamp and flow cell for fast replacement electronic identification of flow cell and lamp with RFID Radio Freq...

Страница 31: ...in which UV absorption takes place and the intensity is converted to an electrical signal by means of the sample photodiode Part of the light is directed to the reference photodiode by the beam split...

Страница 32: ...ID tag reader reads out this information and transfers it to the user interface Figure 16 Flow Cell with RFID tag G 9 iV Table 1 Flow Cell Data STD Semi micro Micro High Pressure Maximum pressure 40 4...

Страница 33: ...an integrated RFID tag that contains the lamp specific information e g part number burn time A RFID tag reader reads out this information and transfers it to the user interface Source Lens Assembly Th...

Страница 34: ...stalled and is processor controlled A photo sensor determines the correct position Mirror Assemblies M1 and M2 The instrument contains two spherical mirrors M1 and M2 The beam adjustable is vertically...

Страница 35: ...the deuterium lamp Beam Splitter Assembly The beam splitter splits the light beam One part goes directly to the sample diode The other part of the light beam goes to the reference diode Photo Diodes A...

Страница 36: ...36 Agilent 1260 Infinity Quaternary LC VL Manual and Quick Reference 1 Introduction Detector...

Страница 37: ...ary LC VL Manual and Quick Reference Agilent Technologies 2 Specifications Physical Specifications 38 Performance Specifications 40 Specification Conditions 47 This chapter provides information about...

Страница 38: ...temperature1 1 This temperature range represents the technical specifications for this instrument The mentioned temperatures may not be suitable for all applications and all types of solvents 4 55 C 3...

Страница 39: ...200 W 683 BTU G1330B Thermostat 20 7 kg 46 lbs 140 x 345 x 435 mm 5 5 x 13 5 x 17 inches 260 VA 210 W 717 BTU G1316B Thermostatted Column Compartment 11 2 kg 25 lbs 140 x 345 x 435 mm 5 5 x 13 5 x 17...

Страница 40: ...0 40 MPa 0 400 bar 0 5880 psi up to 5 mL min Operating range 0 20 MPa 0 200 bar 0 2950 psi up to 10 mL min Pressure pulsation 2 amplitude typically 1 0 or 3 bar at 1 mL min isopropanol at all pressure...

Страница 41: ...cm Warm up cool down time 5 minutes from ambient to 40 C 10 minutes from 40 20 C Dead volume 3 L left heat exchanger 6 L right heat exchanger Communications Controller area network CAN RS 232C APG Re...

Страница 42: ...detection and safe leak handling low voltages in maintenance areas error detection and display Injection range 0 1 100 L in 0 1 L increments recommended 1 L increments Up to 1500 L with multiple draw...

Страница 43: ...accuracy 1 nm After recalibration with deuterium lines Wavelength bunching 2 400 nm Programmable in steps of 1 nm Slit width G4212B 4 nm Fixed slit Diode width 0 5 nm Signal data rate 80 Hz G4212B Sp...

Страница 44: ...reas GLP features Early maintenance feedback EMF for continuous tracking of instrument usage in terms of lamp burn time with user setable limits and feedback messages Electronic records of maintenance...

Страница 45: ...th and 400 bar 5880 psi pressure maximum Micro 2 L volume 3 mm cell path length and 120 bar 1760 psi pressure maximum Semi micro 5 L volume 6 mm cell path length and 40 bar 588 psi pressure maximum Al...

Страница 46: ...output signal for shutdown of pumping system Low voltages in major maintenance areas GLP features Early maintenance feedback EMF for continuous tracking of instrument usage in terms of lamp burn time...

Страница 47: ...V 1 l 2 0 AU 5 typical 2 5 AU 5 ASTM drift tests require a temperature change below 2 C hour 3 6 F hour over one hour period Our published drift specification is based on these conditions Larger ambi...

Страница 48: ...in larger drift Better drift performance depends on better control of the temperature fluctuations To realize the highest performance minimize the frequency and the amplitude of the temperature change...

Страница 49: ...ion Cycle and Low Delay Volume 57 Precise Injection Volume 58 Choice of Rotor Seal 60 Optimizing the Thermostatted Column Compartment 61 Optimizing the Detector Regarding to the System 62 Delay Volume...

Страница 50: ...are empty or changing to solvents that are immiscible with the solvent currently in the vacuum tubes Priming the vacuum degasser by using the quaternary pump at high flow rate is recommended when quat...

Страница 51: ...des A wash bottle filled with water isopropanol 90 10 is placed above the pump in the solvent cabinet and the peristaltic pump moves a flow through the pump head removing all possible buffer crystals...

Страница 52: ...ations and for all gradient analyses For applications using sensitive detectors the compressibility settings can be optimized by using the values for the various solvents described in Table 9 on page...

Страница 53: ...hart Speed 10 cm min 4 Start the recording device with the plot mode 5 Starting with a compressibility setting of 10 10 6 bar increase the value in steps of 10 Re zero the integrator as required The c...

Страница 54: ...ient analysis The residual amount of sample remaining on the outside of the needle after injection may contribute to carry over in some instances When using small injection volumes or when injecting s...

Страница 55: ...d NEEDLE WASH When this command is included in the injector program the needle is lowered once into the specified wash vial before injection For example 1 DRAW 5 l 2 NEEDLE WASH vial 7 3 INJECT Line 1...

Страница 56: ...l use wash vials with an appropriate solvent Using an injector program and several wash vials can be used for cleaning In case the needle seat has got contaminated and carry over is significantly high...

Страница 57: ...ase is directed to the column without passing sample loop needle and needle seat capillary This can help to have faster cycle times especially if low flow rates have to be used like it is mandatory in...

Страница 58: ...her If the mobile phase is not degassed adequately small gas bubbles may form in the sample loop during the injection sequence When using injection volumes 2 L these gas bubbles may affect the injecti...

Страница 59: ...large injection volumes setting the eject speed to a higher value speeds up the injection cycle by shortening the time the metering unit requires to eject solvent at the beginning of the injection cy...

Страница 60: ...mobile phases with pH below 2 3 or above 9 5 or for conditions where the lifetime of the Vespel seal is drastically reduced a seal made of Tefzel is available Tefzel is more resistant than Vespel to...

Страница 61: ...illaries and place them close to the heat exchanger This will reduce heat dissipation and external band broadening Use the left heat exchanger for small volume columns for example 2 3 mm i d columns a...

Страница 62: ...the column Extra Column Volume Extra column volume is a source of peak dispersion that will reduce the resolution of the separation and so should be minimized Smaller diameter columns require proporti...

Страница 63: ...tions describe optimization for signal sensitivity selectivity and linearity spectral sensitivity and resolution DAD only and disk space required for storing data How to Get the Best Detector Performa...

Страница 64: ...ak resolution versus sensitivity versus disk space Use peak width according Choosing a Flow Cell on page 65 as starting point Set the peak width close to the width of a narrow peak of interest in your...

Страница 65: ...tor of about 3 depending on the application Optimizing for Sensitivity Selectivity Linearity and Dispersion Flow Cell Path Length Lambert Beer s law shows a linear relationship between the flow cell p...

Страница 66: ...ht To determine the volume of your peaks take the peak width as reported in the integration results multiply it by the flow rate and divide it by 3 Traditionally LC analysis with UV detectors is based...

Страница 67: ...t in your chromatogram Response time will the be approximately 1 3 of the peak width resulting in less than 5 peak height reduction and less than 5 additional peak dispersion Decreasing the peak width...

Страница 68: ...chromatographic signal Table 11 Peak Width Response Time Data Rate Peak width at half height min 1 1 Values in the User Interface may be rounded Response s Signal data rate Hz Scan data rate HZ 126 pt...

Страница 69: ...254 0 4 reference 360 0 100 that is the average absorbance from 252 256 nm minus the average absorbance from 310 410 nm As all analytes show higher absorbance at 252 256 nm than at 310 410 nm this sig...

Страница 70: ...averaging over a wavelength range compared to a 4 nm bandwidth the baseline noise is reduced by a factor of approximately 2 5 whereas the signal is about 75 of a 4 nm wide band The signal to noise ra...

Страница 71: ...ctive index changes during a gradient An example of the reduction of baseline drifts is shown in Figure 21 on page 71 for PTH amino acids Without a reference wavelength the chromatogram drifts downwar...

Страница 72: ...ibrary searches Reducing the spectrum storage range saves disk space Step Most substances have broad absorbance bands Display of spectra peak purity and library search works best if a spectrum contain...

Страница 73: ...an example the analysis of hydrochlorothiazide in the presence of caffeine is described If hydrochlorothiazide is analyzed in biological samples there is always a risk that caffeine is present which...

Страница 74: ...much as 10 30 Figure 23 Peak Suppression Using Reference Wavelength Ratio Qualifiers for Selective Detection of Compound Classes Ratio qualifiers can be used where in a complex sample only one partic...

Страница 75: ...qualifier criterion and therefore the output was set to zero The characteristic wavelengths 249 nm 1 and 224 nm 2 were found from the spectra shown in Figure 24 on page 75 The ratio range was set at...

Страница 76: ...is used as self test of the heater functionality 1 to 30 minutes the heater control is OFF and the heater element runs at 40 duty cycle After 30 minutes the heater control is ON and is working with op...

Страница 77: ...inity Quaternary LC VL Manual and Quick Reference 77 Optimization of the Agilent 1260 Infinity Quaternary LC VL 3 Optimizing Detection with DAD Figure 26 Detector Warm up 1st hour Figure 27 Detector W...

Страница 78: ...or Performance The detector has a variety of parameters that can be used to optimize performance The information below will guide you on how to get the best detector performance Follow these rules as...

Страница 79: ...h the cell volume usually increases in the example from 5 13 L Typically this causes more peak dispersion As demonstrated this did not affect the resolution in the gradient separation in the example s...

Страница 80: ...tion on path lengths of the VWD flow cells The correct response is expected response correction factor Please find below the details of the Agilent 1200 Infinity Series Variable Wavelength Detector fl...

Страница 81: ...cut off wavelength of the mobile phase at a wavelength where the analytes have strong absorptivity if you want to get the lowest possible detection limit at a wavelength with moderate absorptivity if...

Страница 82: ...82 Agilent 1260 Infinity Quaternary LC VL Manual and Quick Reference 3 Optimization of the Agilent 1260 Infinity Quaternary LC VL Optimizing Detection with VWD...

Страница 83: ...lent Technologies 4 System Setup and Installation Installing Software 84 Installing the Modules 85 Priming the System 86 Integration Into the Network 90 This chapter includes information on software i...

Страница 84: ...ductivity through automatic notification of maintenance and service needs with the use of advanced counters This allows a problem to be fixed before it impacts results The software includes an extensi...

Страница 85: ...es for the modules refer to the individual module manuals These manuals also contain information on specifications maintenance and parts Stack Configuration You can ensure optimum performance by insta...

Страница 86: ...lvents and reagents can bear health risks Please observe appropriate safety procedures for example goggles safety gloves and protective clothing as described in the material handling and safety data s...

Страница 87: ...e tubing of channel A1 is advancing towards the pump If it isn t disconnect the solvent tubing from the solvent selection valve attach a syringe with a syringe adapter and pull the liquid through the...

Страница 88: ...omposition and flow rate for your application and close the purge valve 5 Pump for approximately 10 minutes before starting your application When When the pumping system has been turned off for a cert...

Страница 89: ...t steps 1 to 7 for the other channel s of the pump 9 Install the desired column set the required composition and flow rate for your application and equilibrate the system for approx 10 minutes prior t...

Страница 90: ...itching between reverse phase and normal phase both times Isopropanol Isopropanol Best solvent to flush air out of the system Miscible with almost all solvents After an installation Ethanol or methano...

Страница 91: ...Quick Start Guide Preparing the System 92 Turning the System ON 92 Loading the Default Method 93 Configuring the Online Plot 94 Purging the Pump 96 Setting Up the Method 97 This chapter provides infor...

Страница 92: ...rt All Programs Agilent ChemStation Agilent ChemStation Instrument 1 Online As more than one instrument system may be connected to the computer the number 1 2 indicates the system number The ChemStati...

Страница 93: ...hod 1 Go to Method and Run Control view of the ChemStation 2 On the menu bar select Method New Method and select DEF_LC M from the context menu Alternatively you can use the Load Method icon under the...

Страница 94: ...1 If the Online Plot window is not visible Click View Online Signals Signal Window 1 to display the window 2 To configure the desired signal s in the Online Plot window click Change The Edit Signal Pl...

Страница 95: ...ng at previous injections The X and Y axis scales can be adjusted directly with the up down buttons on each axis The Adjust button in the Online Plot window moves the current point on the selected sig...

Страница 96: ...idle for a few hours or more air may have diffused into the solvent lines and purging is recommended The solvent reservoirs are refilled and the pump requires purging to fill the system with fresh sol...

Страница 97: ...nfinity Quaternary Pump enter the following parameters Flow rate 1 5 ml min Solvent A Select Water from the compressibility drop down list Solvent B Select the check box to make Solvent B active B Ini...

Страница 98: ...Infinity DAD enter the following parameters Use Signal Turn all signals except Signal A off by clearing the check boxes Signal A 250 nm bw 100 nm ref 360 nm bw 100 nm Peak width 0 012 min 0 25 s Resp...

Страница 99: ...l and Electronic Equipment Directive 103 Lithium Batteries Information 104 Radio Interference 105 Sound Emission 106 Solvent Information 107 UV Radiation 108 Declaration of Conformity for HOX2 Filter...

Страница 100: ...dicates eye damage may result from directly viewing the light produced by the deuterium lamp used in this product The apparatus is marked with this symbol when hot surfaces are available and the user...

Страница 101: ...the following must be observed Do not remove instrument covers when operating Before the instrument is switched on all protective earth terminals extension cords auto transformers and devices connect...

Страница 102: ...components with power cable connected Do not operate the instrument in the presence of flammable gases or fumes Operation of any electrical instrument in such an environment constitutes a definite sa...

Страница 103: ...and electronic appliances starting with 13 August 2005 NOTE This product complies with the WEEE Directive 2002 96 EC marking requirements The affixed label indicates that you must not discard this ele...

Страница 104: ...e disposed off locally according to national waste disposal regulations for batteries Replace only with the same or equivalent type recommended by the equipment manufacturer WARNING Lithiumbatteri Eks...

Страница 105: ...d to provide optimized protection against radio interference All cables are in compliance with safety or EMC regulations Test and Measurement If test and measurement equipment is operated with unscree...

Страница 106: ...r s Declaration This statement is provided to comply with the requirements of the German Sound Emission Directive of 18 January 1991 This product has a sound pressure emission at the operator position...

Страница 107: ...igh concentrations of inorganic acids like sulfuric acid and nitric acid especially at higher temperatures if your chromatography method allows replace by phosphoric acid or phosphate buffer which are...

Страница 108: ...personnel is limited to the following TLVs Threshold Limit Values according to the American Conference of Governmental Industrial Hygienists Typically the radiation values are much smaller than these...

Страница 109: ...Agilent 1260 Infinity Quaternary LC VL Manual and Quick Reference 109 Appendix 6 Declaration of Conformity for HOX2 Filter Declaration of Conformity for HOX2 Filter...

Страница 110: ...ent Technologies on Internet Agilent Technologies on Internet For the latest information on products and services visit our worldwide web site on the Internet at http www agilent com Select Products C...

Страница 111: ...online plot 94 control and data evaluation 43 45 cool down time 41 correction factors for flow cells 66 80 cutoff filter 34 D data evaluation and control 43 data rate sampling rate 45 dead volume 41 d...

Страница 112: ...re 38 operational hints vacuum degasser 50 operational pressure range 40 optical unit flow cell 32 optical unit beam splitter assembly 35 entrance slit assembly 33 filter assembly 34 filter 34 grating...

Страница 113: ...seal alternative material 52 selectivity optimization 73 setable flow range 40 slit width 43 slit fix 28 solvents 107 sound emission 106 source lens assembly 33 specifications analog signal output 44...

Страница 114: ...0310 www agilent com Agilent Technologies In This Book This manual contains technical reference information about the Agilent 1260 Infinity Quaternary LC VL The manual describes the following introduc...

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