Mass Spectrometry Laboratory and Services
The Mass Spectrometry facility in Chemistry Department at Rutgers University Newark Campus provides high end mass spectrometry services for the Rutgers University Community. We have liquid chromatography system coupled with high resolution mass spectrometer to provide separation and analysis of complex mixtures.
We are located in the Life Science Center, room 001, at 225 University Avenue, and Olson Hall, room 006, at 73 Warren Street, Newark, NJ.
The facility manager Dr. Roman Brukh is available to discuss your specific experimental needs.
Our services include:
High resolution mass measurements, which can provide elemental composition for analytes.
Electrospray ionization (ESI), Atmospheric Pressure Chemical ionization (APCI), and Matrix Assisted Laser Desorption Ionization (MALDI) mass spectrometry are used for wide range of applications by research groups and students.
Fragmentation methods include in-source collision activated dissociation (IS-CAD), collision cell activated dissociation (C-CAD), sustained off-resonance irradiation collision activated dissociation (SORI-CAD), infrared multi-photon dissociation (IRMPD), and electron capture dissociation (ECD).
High Performance Liquid Chromatography is coupled with ESI or APCI sources to allow for the separation of mixtures and subsequent analysis.
Sample applications are compound elucidation, drug impurity analysis, natural product characterization, high-end proteomics studies, polymer and metabolomics research.
The mass spectrometry facility has three mass spectrometers available:
1. Bruker FTMS equipped with a 7.0 T magnet, and an Apollo II Dual ESI/MALDI source that can provide simultaneous operation in both modes.
The HPLC system is available to be coupled with ESI/APCI source for separation and analysis of complex mixtures.
MS/MS or MS/MS/MS fragmentation data provides a tremendous amount of information for analysis of unknown samples.
2. Bruker UltrafleXtreme MALDI-TOF/TOF is used for a variety of MALDI applications, including protein identification, peptide fingerprinting, polymer research, and structure identification for a wide spectrum synthetic and biological compounds.
3. Agilent 6890 GC equipped with 5973 MSD GC-MS system provides routine mass spectrometry analysis for our graduate and undergraduate students.
Fourier Transform Mass Spectrometry
The FTMS is equipped with a 7.0 T actively shielded, high-stability magnet, and an Apollo II Dual ESI/MALDI source to provides simultaneous operation in both modes. The ESI source has an ion funnel and a spray chamber with a grounded off-axis sprayer. The MALDI source has computer controlled X-Y stage for scout-385 MTP target plates. Additional instrument capabilities are HPLC and nanospray experiments. Fragmentation methods include in-source collision activated dissociation (IS-CAD), collision cell activated dissociation (C-CAD), sustained off-resonance irradiation collision activated dissociation (SORI-CAD), infrared multi-photon dissociation (IRMPD), and electron capture dissociation (ECD).
Sample applications are compound elucidation, drug impurity analysis, natural product characterization, high-end proteomics studies, and metabolomics research.
Electrospray Ionization (ESI) Instructions
The solvent should ensure that the analyte is susceptible to charging. Acid in concentration of 0.1% is often added. Solution must be conductive.
For proteins: 1:1 water and methanol + 0.1% formic or acetic acid.
ESI compatible solvents: water, acetonitrile, methanol, dichloromethane, DMSO, isopropanol, butanol, THF, acetone, DMF.
ESI compatible buffers: acetic acid, formic acid, ammonium acetate, ammonium hydroxide.
Do not work well for ESI: hydrocarbons, aromatics, carbon tetrachloride, toluene.
Deionized water with conductivity >18 Ω/cm3 should be used. Impurities in organic solvents can result in unwanted peaks.
For example, sodium, Na, can be introduced even from glassware. Sulfate and phosphate impurities in protein samples can lead to an adduct with mass of 98 m/z. Common surfactants, such as sodium dodecyl sulfate (SDS) and Triton X in protein samples can completely mask the protein signal.
The sample concentration should be in the range of 0.5-5 μM. The sample should be clear. Microscopic debris can clog the electrospray needle.
Getting an ESI Signal
Login to your account. Write the information needed in the Log Book. Open "apexControl Standard" software.
Load the sample or standard into the 100 μL or 250 μL syringe. Set the pump flow to 120-300 μL/hr, connect the syringe to the injection port and turn on the flow. Fast forward the sample through the tubing to remove any air in the system. Load calibrated ESI method or open a previously acquired file. This will load a set of instrumental parameters. Make sure that there is capillary current (15-20 μA) present in the "Readback" tab. If there is no current, check if the "Nebulizer Gas Flow", "Drying Gas Flow", "ESI High Voltage", and the "Drying Gas Heater" are turned on. Set the number of scans to average 10-50.
The "Tune" mode allows the operator to optimize signal.
In order to acquire a data file, edit the "Prefix" field and the "Subdirectory" field. Click "Acquisition/Run Method" to acquire the spectrum.
Once a spectrum has been acquired, the FT-ICR must be calibrated.
Open the correct mass list. Select the calibration mode - Cal2 or Cal3. Select the peak to be calibrated in the "Current Mass" column of the corresponding reference mass. For Cal2, a minimum of 3 peaks are required. For Cal3 - minimum 4 peaks. Click "Accept". To check this calibration, click "Automatic". Repeat this process until the entire mass range is calibrated.
Fourier Transform Mass Spectrometry
The FTMS offers the following ion dissociation methods for MS/MS experiments:
Matrix Assisted Laser Dissorption Ionization - Time of Flight
The Ultraflex is a MALDI (Matrix Assisted Laser Desorption/Ionization) tandem mass spectrometer specially designed for automated MS and MS/MS analysis. Tandem Mass Spectrometry is a technique that utilizes more than one mass selective stage in a mass spectrometer. The most common form in practice is a two stage arrangement to record MS/MS spectra. The incorporated LIFT device allows acquiring full fragment ion spectra within one single scan and replaces traditional measurements of segmented spectra with stepwise-reduced reflector potentials.
α-Cyano-4-hydroxycinnamic acid (α-Cyano; HCCA; CCA)
Commonly used for peptides in the lower mass range.
Not soluble in water and well soluble in organic solvents.
It is considered a "hard" matrix, which means the analyte molecules get a lot of internal energy during desorption and ionization. This leads to a considerable amount of ion fragmentation (post source decay - PSD). If peptides of small molecular weight are measured and the laser power is chosen only slightly above the threshold, this is not a problem. If the analyte molecules become bigger, however, the probability of the fragmentation increases until almost all of the analyte ions undergo fragmentation. Therefore α-Cyano is the matrix of choice for PSD-analysis.
The main advantage of α-Cyano in the measurement of peptides is the ability of this matrix to form small homogenous crystals. Since geometric inhomogeneity relates directly to decreased resolution in the MALDI-analysis, α-Cyano preparations usually yield good resolution.
Sinapinic Acid (SA)
Sinapinic Acid is most commonly used in the analysis of high mass proteins.
Not soluble in water but well soluble in organic solvents.
Compared to α-Cyano it is a "softer" matrix. The analyte Ions get less internal energy and the amount of fragmentation is smaller, making this matrix more suitable for measurement of proteins. Sinapinic Acid also can form small crystals. However, Sinapinic Acid tends to form adducts with the analyte ions. These adducts can be resolved in the mass spectrum for proteins up to 40 kD.
2,5-Dihydroxybenzoic acid (DHB)
This is the matrix of choice for the preparation of glycoproteins and glycans. It is also often used for peptides.
Unlike α-Cyano and Sinapinic Acid it is soluble in water as well as organic solvents.
The main disadvantage of DHB is the fact that it forms big crystal needles. This means that the geometry of the sample changes from spot to spot. If spectra are summed up from different spots on the sample, the resolution is considerably lower than spectra obtained from an α-Cyano.
On a steel target, DHB preparations will form a crystalline ring. Good peptide spectra are usually only obtainable at the rim.
The main advantage of DHB for MALDI of peptides is the fact that this matrix is more tolerant towards contaminations.
MALDI Sample Preparation
The ideal sample preparation in MALDI would be a homogenous layer of small matrix crystals containing a solid solution of the analyte.
To obtain the best result, there is a choice of different matrices as well as preparation techniques. The choices depend on the nature of the analyte.
One aim is to obtain a homogenous preparation of the matrix, both in terms of sample distribution and in term of the sample geometry.
Preparing the sample on the target
Like the choice of the matrix compound, there is also a choice of how to actually prepare the sample. This section discusses the conventional targets. Anchor-targets have to be prepared using specialized anchor-chip protocols (refer to the anchor chip manual).
The chemicals should be of highest available purity.
Saturated matrix solutions should be prepared freshly.
Dried droplet method
A saturated matrix solution is prepared. Unless special solbents have to be used, the solvent used is TA (33% Acetonitrile, 0.1% TFA).
This matrix solution is mixed in equal volumes with the sample solution. The mixture is pipeted on the target (0.5 to 1 µl) and dried at ambient temperature.
The preparation might yield relatively large crystals on the target surface.
The advantages of this method are:
- the method is suitable if the sample contains organic solvents;
- if a "sweet spot" is found on the preparation, a large number of laser shots can be applied to that spot;
- if the sample contains contaminants, there is a chance, that analyte and contaminants will crystallize at spatially different regions on the target;
- the sample can be washed after the crystallization to remove salts;
- the sample can also be recrystallized after washing.
Disadvantages include the need to search for sweet spots and the limited resolution due to the large crystals.
Gas Chromatography Mass Spectrometry
Agilent 6890 GC equipped with 5973 MSD GC-MS system provides routine mass spectrometry analysis for our graduate and undergraduate students.
High Performance Liquid Chromatography
The Agilent HPLC system is available to be coupled with ESI/APCI source for separation and analysis of complex mixtures.
The mass spectrometry laboratory performs several types of services including training of new students. The most common service is nominal and accurate mass measurements.
Electrospray Ionization (ESI)
The ESI source is used for the measurements of singly charged samples (small molecules) and multiple charged samples such as proteins, and peptides. The sample solution is introduced through the nebulizer assembly into the spray chamber, where it is subjected to the ESI process by means of an electrical field between the inner chamber wall and the spray shield, and with the aid of a nebulizer gas (nitrogen). Heated drying gas, flowing in the opposite direction of the stream of droplets is used to aid volatilization and ionization, and to carry away any uncharged material. The source assembly delivers the pressurized drying gas and guides it past the spray shield into the spray chamber. Ions are attracted by the electrical field strength between the spray chamber (ground potential) and the negatively biased metal-coated glass capillary, the inlet to the vacuum system (positive mode). A potential difference of about 400 V between the spray shield and the tip of the glass capillary with the spray shield at a lower voltage acts as a further ion pull into the vacuum system.
Atmospheric Pressure Chemical Ionization (APCI)
The APCI source is best used for the analysis of polar and non-polar analytes. The nebulization process for this ion source is similar to that of the ESI source. However, APCI nebulization takes place in a heated vaporizer tube. The heat evaporates the spray droplets resulting in gas-phase solvent and sample molecules. On leaving the vaporizer tube, gas phase solvent molecules are ionized by a current regulated discharge from a corona needle at a voltage of 1-4 kV. By transferring their charge, the solvent ions convert sample molecules to sample ions.
Matrix Assisted Laser Desorption Ionization (MALDI)
A MALDI sample is prepared by mixing an analyte with a suitable matrix compound on a metal sample plate. Evaporation of the solvent causes a co-crystallization process of both matrix and analyte material. The incorporation of the sample molecules into the lattice structure of the matrix is a pre-condition for a successful laser desorption/ionization process. The crystallized surface of the sample-matrix mixture is then exposed to an intensive pulse of short-wave laser irradiation. The photo-ionized radical matrix molecules cause a high yield of electrically charged sample molecules by transferring protons.
Electron Ionization (EI)
In electron ionization process, an electron from the analyte molecule is expelled during the collision of a molecule with highly energetic (70 eV) bombarding electrons produced by a heated filament. The radical cation products are then directed towards the mass analyzer. Due to the high energy of bombarding electrons some bond dissociation reactions can be observed. These ions are known as second-generation product ions. The ionization process often follows predictable cleavage reactions that give rise to fragment ions which convey structural information about the analyte.
Nominal mass of a compound is the mass of the most abundant constituent element isotope of each elements rounded to the nearest integer value and multiplied by the number of atoms of each element.
Nominal mass cannot be used to calculate the elemental composition of a compound.
All our mass spectrometers can be used to measure nominal mass.
Accurate mass measurements
Accurate mass measurements are done using our FTMS and MALDI-TOF mass spectrometers.
The typical mass accuracy is within 2 ppm but for many compounds and methods we can achieve accuracy of less than 1 ppm.
Accurate mass can be used for calculation of the elemental composition of a compound, and together with the simulated isotopic pattern it is used for the compound identification.
Routine Services for Rutgers University
Low resolution - $10
High resolution - $30
Advance analysis including LCMS - $75/hour
MALDI - $20
Routine Services for Academics and Non Profit
Low resolution - $15
High resolution - $50
Advance analysis including LCMS - $100/hour
Routine Services for Profit
Low resolution - $40
High resolution - $100
Advance analysis including LCMS - $150/hour
For information on other services and sample submission please e-mail us email@example.com
Full text, PDF format, Rutgers restricted
Mass Spectrometry: A Foundation Course. Kevin Downard.
Mass Spectrometry: A Textbook. Jürgen H. Gross.
The Mass Spectrometry Laboratory at Chemical Department Rutgers-Newark is located in the Life Science Center, room 001, at 225 University Avenue, and Olson Hall, room 006, at 73 Warren Street, Newark, NJ.
Please contact us at: