Mass Spectrometry Facility

The Orbitrap MS was acquired through an NSF MRI MS grant obtained by Co-PIs Phil Crews and Ted Holman and was installed in 2014. It brings the first instrument to the campus possessing high mass accuracy in full ion scan, and MSn modes accompanied by substantial resolving power. The purpose of obtaining this equipment for the analysis of small molecules was to bring cutting edge experiments and data to our class rooms, and open up new avenues in UCSC programs by using MS to recruit and train students and postdocs.

LC-MS

Liquid chromatography is a powerful technique for analyzing complex mixtures. Detection of compounds in mass spectrometry relies on the ability to ionize target compounds. In complex mixtures, low abundance compounds or those that are difficult to ionize may be masked by higher abundance, easily ionized compounds. This type of interference is referred to as “suppression”. Suppression effects can be reduced or eliminated by separating the compounds to be analyzed before they are introduced into the mass spectrometer. We have a Thermo LTQ-MS and a Thermo Velos-Pro Orbitrap MS instrument to use with a variety of columns and solvent systems to screen for compounds of interest. We also frequently use this instrument for analysis of natural products mixtures, to follow the progress of synthetic reactions and to analyze constituents of other bio-materials such as fuels or fluids.

MSn for Structural Analysis

The structure of a compound can be deduced by fragmenting the parent ion and measuring the masses of the fragment ions. The MS facility has a Thermo LTQ-MS and an LTQ-Orbitrap Velos Pro-MS. The LTQ is an electrospray ionization (ESI) mass spectrometer with a linear ion trap analyzerand a mass range of up to 4000. The LTQ can be set up to collect full scan MS and MSn (n=2-10) data. Advanced scanning modes include data-dependent scans, zoom scans, and ultra zoom scans.The LTQ is designed with interchangeable probes of electrospray ionization and atmospheric pressure chemical ionization (APCI). The Orbitrap can iteratively isolate parent ions and measure the masses of fragments produced by collision induced dissociation (CID), pulsed Q collision-induced dissociation (PQD) and high energy collision induced dissociation (HCD, Orbitrap only). Thus we can sequentially break the parent compound into smaller and smaller pieces to learn about its structure.

Accurate Mass Analysis

When characterizing novel compounds a mass measurement with sufficient resolution and mass accuracy can be used to generate a molecular formula. Publication of new compounds in ACS journals requires MS accuracy data at the level of 3 amu. Our Orbitrap mass spectrometer is capable of mass measurements with sub-ppm mass accuracy and resolution (FWHM) in the range of 4,000 amu. This is sufficient for assigning formulae for most compounds up to ~500 amu.

Proteomics Mass Analysis

LTQ is designed to assist with proteomics type studies. For example, proteins are digested with trypsin and the resulting peptides are analyzed by LC/MS/MS. The peptides are fragmented in the mass spectrometer to yield diagnostic patterns that can be matched to protein sequence databases via computer algorithms. The sensitivity of this technique is 1 ng or less starting material for a given protein.

Please contact Professor Ted Holman for current fee schedule at holman@ucsc.edu.

To use the MS Facility, please make a reservation at the website https://faces.ccrc.uga.edu/

Required Procedures

1. DO keep sample amounts low. For most applications on the LTQ instruments sensitivity is on the order of pmols. Keep concentrations in the low hundreds of nanograms per milliliter. If you are working with a new sample and don’t know how it will behave, start with a very low concentration and go higher until you see a signal.
2. DO make the sample as clean as possible. Purity and amount are the two big determinants for getting good mass spectrometry results.
3. DO work with volatile solvents and buffers. Volatile buffers will pass through the mass spectrometer. Non-volatile buffers, salts etc., leave residue in the source, in the capillary, and in the mass spectrometer necessitating costly, time-consuming clean outs.
4. DO ask questions. Li and Postdoc advisers are happy to consult with all the users regarding any and all aspects of mass spectrometry. We are glad to help with or consult about the experiments, since cleaner samples means higher quality data for all the users. It is always better to ask than risk damaging columns or instruments by using inappropriate techniques or reagents.
5. DO centrifuge your samples before loading them in the vials for mass spectrometric analysis. Particulates clog columns leading to high back pressures. Filtration is another option, but centrifugation is preferred.

Forbidden Procedures

1. DO NOT operate the instruments without appropriate training. Training for the walk-up is offered frequently by appointment. If you require mass spectrometry for your project and you have not been trained, ask another member of your lab or another lab to run the samples for you until you complete the training.
2. DO NOT use trifluoroacetic acid (TFA). While TFA is an excellent pairing reagent and gives nice peak shape for HPLC, it dramatically reduces ionization of most compounds at the source.
3. DO NOT try to analyze samples with high concentrations of mineral salts (Na, K etc.). Excess non-volatile buffer components can be deposited on the source during ionization and reduce sensitivity. They can also build up in other parts of the instrument and cause problems.
4. DO NOT spike samples with Na or K.
5. DO NOT analyze samples with high concentrations of solvents/conditions (e.g. hexanes and high pH) that are not recommended in the column care guide. You could damage or dissolve the matrix, ruining the column.
6. DO NOT analyze samples with corrosive compounds present. Perchloric acid and other corrosives will damage or destroy the stainless steel parts of the HPLC system.
7. DO NOT attempt to fix or recalibrate the instruments without contacting the facility managers or Postdoc Super Users. Incorrectly calibrating or tuning the instrument will affect data collection in all experiments after you. If you believe the instrument is not working correctly, please report the problem.

If you are not yet trained to use the equipment, you may complete our Request a Training Form.

Please send email to Qiangli Zhang (qzhang@ucsc.edu). Facility manager, Li Zhang, is only 50% time. Running of submitted samples will be on a staff time availability basis.

To report a problem, please email to Qiangli Zhang (qzhang@ucsc.edu).