Navigation Links
Detection of Alprazolam and Temazepam in a Bovine Plasma Matrix Using the Varian 500-MS Ion Trap Mass Spectrometer

Bethany V. Pond, Jason S. Wood, and August Specht Varian, Inc.

Introduction

Alprazolam (Xanax) and Temazepam (Restoril) were analyzed in a Bovine Serum Albumin (BSA) matrix, both before and after protein precipitation using Liquid Chromatography Electrospray Ionization Mass Spectrometry (LC/ESI/MS/MS). The ability of the Varian 500-MS IT Mass Spectrometer to detect and accurately quantitate these drugs in the BSA matrix is investigated here.

Instrumentation

Varian ProStar 210 Solvent Delivery Module (2)

Varian 500-MS Ion Trap Mass Spectrometer equipped with an ESI source

Varian Prostar 420 Autosampler

Materials and Reagents

All solvents (reagent or HPLC Grade) and the Bovine Serum Albumin (Part # A4503-10G) were purchased from Sigma-Aldrich Corporation (St. Louis, MO). The analytes, Temazepam (Part # T-907) , Alprazolam (Part # A-903), and Alprazolam-d5 (Internal Standard, Part #A-910), were purchased from Cerilliant Corporation (Round Rock, TX).

Sample Preparation

Stock solutions were purchased at a concentration of 1 mg/mL in methanol. Further dilutions of the stock solutions were carried out in a 1:1 mixture of 2mM ammonium acetate in 0.1 % aqueous formic acid: 0.1% formic acid in acetonitrile.

No Matrix Sample Preparations:

Solutions ranging in concentration from 0.1-1000 pg/μL of alprazolam and temazepam were made from the stock solution.

Post-Protein Crash Matrix Preparation:

Equal Volumes of 0.3 mM Bovine Serum Albumin (BSA) in Phosphate Buffered Saline (PBS) and an 80:20 Acetonitrile:Methanol solution containing 0.1% Acetic Acid were centrifuged for 5 minutes at 12000 rpm and refrigerated for 1.5 hours. The supernatant was removed and spiked with the internal standard (10 pg/μL of Alprazolam-d5) and the analytes in the same concentrations as used in the no matrix preparation

Pre-Protein Crash Matrix Preparations:

0.3 mM BSA in PBS was spiked with the internal standard (10 pg/μL of Alprazolam-d5) and the analytes in the same concentrations as the no matrix preparations. An equal volume of the 80:20 acetonitrile:methanol solution containing 0.1% acetic acid was added to the spiked BSA solutions. The mixtures were centrifuged for 10 minutes at 12000 rpm and then refrigerated for 1.5 hours. The supernatant was removed and subsequently analyzed.

The samples were run in triplicate for calibration curve purposes on a 500-MS Ion Trap Mass Spectrometer.

Each analyte was run individually with the internal standard in an MRM experiment for a total run time of 5 minutes. For the no matrix run, the analytes and internal standard were run individually in a MS/MS experiment with the product scan centered around the specific product ions.

Results and Discussion

Calibration curves were run without the matrix, with theafter crash spiked matrix, and the before crash spiked matrix with good linearity observed for each analyte. The extracted ion chromatogram of 500 fg (on column) of Temazepam (m/z = 255.2) in the before crash matrix is shown in the top panel of Figure 1, while the bottom panel of Figure 1 is the extracted ion chromatogram of the internal standard, Alprazolam-d5 (50 pg on column). The Signal-to-Noise (S/N P-P) ratio of temazepam in the spiked matrix was determined to be 16:1 at 500 fg on column.

The LOD (limit of detection) values for the three different types of matrix environments are summarized in Table 1. A calibration curve of Alprazolam in a before crash spiked matrix of these compounds is shown in Figure 2. An R2 value > 0.99 was observed for both compounds in all of the matrices studied, over the calibration range of 0.2 pg/μL 1000 pg/μL (1 pg 5000 pg on column). An LOQ = 0.2 pg/μL (1 pg on column) was determined for all three matrix conditions for both Alprazolam and Temazepam.

Even with the addition of the matrix, the range of quantitation did not change. Figure 3 shows the extracted ion chromatogram (m/z = 255.2) of Temazepam in all three of the matrix conditions.

It is evident from Figure 3 that the 500-MS Ion Trap is capable of handling a biological matrix and thus is still able to reach a low level of detection under the conditions described. These results show that the 500-MS Ion Trap is a sensitive and quantitative tool for the detection of target analytes in biological matrices.

Conclusion

The measurements taken with Alprazolam and Temazepam spiked into bovine plasma matrix illustrate that it is possible to reach low levels of quantitation (low- to sub-pg level) using the 500-MS Ion Trap Mass Spectrometer. Quantitative analysis of Temazepam and Alprazolam in a biological matrix has been demonstrated in the 500-MS instrument illustrating its excellent performance in both quantitative and qualitative analysis.


'"/>

Source:


Page: All 1 2 3

Related biology technology :

1. Simple, Sensitive, and Rapid Detection of FLAG -Tagged Fusion Proteins
2. A New PCR-based Mycoplasma Detection Method
3. HSVision Molecular Beacon Detection Module Rapidly Detects Herpes Simplex Virus DNA
4. Detection and Identification of Phosphorylation Sites in Proteins Using LC/MS/MS with Neutral Fragment Loss Mapping
5. Detection of mRNAs on Cryosections of the Cardiovascular System Using DIG-Labeled RNA Probes
6. Gene Expression Arrays: Highly Sensitive Detection of Expression Patterns with Improved Tools for Target Amplification
7. The DIG System Nonradioactive and Highly Sensitive Detection of Nucleic Acids
8. Quantitative Measurement of Cell Proliferation Using the BrdU ELISA: A Comparison Between Colorimetric and Chemiluminescent Detection
9. Quantification of Nucleosomes in Serum by the Cell Death Detection ELISAplus
10. A Further Step in Understanding Apoptosis Direct Detection of PARP Cleavage
11. In Situ Cell Death Detection Kit
Post Your Comments:
(Date:7/31/2015)... Dallas, Texas (PRWEB) , ... July 31, 2015 ... ... research report, offering competitive market analysis and premium industry insights on the global ... challenges, based on a collective study of vivid market scenarios and analysis of ...
(Date:7/31/2015)... ... 31, 2015 , ... R-Biopharm is proud to announce the ... in foods, has been accepted by AOAC International as Official First Action method. ... Cereal-Based Products by R5 Competitive ELISA,” based on a specific monoclonal antibody to ...
(Date:7/30/2015)... 30, 2015 AACC welcomed thousands of ... AACC Annual Meeting & Clinical Lab Expo in ... The meeting showcased revolutionary advancements in clinical testing ... healthcare providers to diagnose patients quickly and accurately ... As of Wednesday, July 29, ...
(Date:7/30/2015)... ... July 30, 2015 , ... Tronics, a ... Sales & Marketing Director. , With more than 25 years of experience ... reinforcing Tronics’ business development activities worldwide. He brings to the company his strong ...
Breaking Biology Technology:Resorcinol Market to Grow at 6% CAGR to 2019 Says New Research Report at ReportsnReports.com 2Resorcinol Market to Grow at 6% CAGR to 2019 Says New Research Report at ReportsnReports.com 3Resorcinol Market to Grow at 6% CAGR to 2019 Says New Research Report at ReportsnReports.com 4RIDASCREEN® Competitve Gliadin Receives AOAC Official First Action Status 2Innovative Research on Cancer, Heart Disease, and Infectious Diseases Draws More than 17,500 Attendees to 2015 AACC Annual Meeting 2Innovative Research on Cancer, Heart Disease, and Infectious Diseases Draws More than 17,500 Attendees to 2015 AACC Annual Meeting 3Innovative Research on Cancer, Heart Disease, and Infectious Diseases Draws More than 17,500 Attendees to 2015 AACC Annual Meeting 4Tronics appoints Francois Vieillard as Sales and Marketing Director 2
... Public Health and Safety is a major ... homes and everywhere in between, dangerous germs like MRSA and ... and more people are contracting various strains of bacteria and ... approximately 15 to 60 million people contract seasonal flu each ...
... BioSpecifics Technologies Corp. (Nasdaq: BSTC ), a ... announced that BioSpecifics, President, Tom Wegman, will present at ... on Wednesday, September 15, 2010, at 9:35am EDT. ... be accessed under "Calendar of Events" in the Investor ...
... FRANCISCO and ILLKIRCH, France, Sept. 8 Jennerex, ... the development and commercialization of first-in-class targeted oncolytic ... FR0005175080), a bio-pharmaceutical company specialized in the development ... entered into an exclusive partnership to develop and ...
Cached Biology Technology:SpectraSan 24 Holds First Public Health and Safety Symposium in Ohio 2SpectraSan 24 Holds First Public Health and Safety Symposium in Ohio 3Jennerex and Transgene Enter Into an Exclusive Partnership for the Development and Commercialization of JX-594 for the Treatment of Cancers 2Jennerex and Transgene Enter Into an Exclusive Partnership for the Development and Commercialization of JX-594 for the Treatment of Cancers 3Jennerex and Transgene Enter Into an Exclusive Partnership for the Development and Commercialization of JX-594 for the Treatment of Cancers 4Jennerex and Transgene Enter Into an Exclusive Partnership for the Development and Commercialization of JX-594 for the Treatment of Cancers 5
(Date:7/8/2015)... , N.J. and NEW YORK , July ... Guidepoint today announced BD & Guidepoint ... companies with free access to Guidepoint,s expert network services. ... that are developing cutting-edge technologies to improve healthcare delivery ... Guidepoint research manager, each start-up entrepreneur will be able ...
(Date:7/7/2015)... Conn. , July 7, 2015 NXT-ID, Inc. (NASDAQ: ... focused on the growing mobile commerce market announces a revised version ... this week. The commercial will air on CNBC in ... Los Angeles and San Francisco ... The commercial focuses on Wocket,s ability to replace all the cards ...
(Date:6/30/2015)... Pa. , June 30, 2015 ... the company as CEO to help further develop Genisphere,s ... wealth of partnering experience, having spent much of the ... including Therakine, Amgen, Baxter Bioscience, and Johnson & Johnson. ... focus on Technology and Biotech Corporate Finance. He graduated ...
Breaking Biology News(10 mins):BD and Guidepoint Team to Connect Healthcare Start-up Companies With Business and Scientific Experts 2BD and Guidepoint Team to Connect Healthcare Start-up Companies With Business and Scientific Experts 3Wocket Smart Wallet Commercials Airing on CNBC to Showcase This Summer's Must- Have Tech Product 2Wocket Smart Wallet Commercials Airing on CNBC to Showcase This Summer's Must- Have Tech Product 3Genisphere Appoints New CEO, Thomas Bliss 2Genisphere Appoints New CEO, Thomas Bliss 3
... Iowa State University researchers have developed a technology that ... cervical cancer in women. Thats a significant improvement ... Edward Yeung, an Iowa State Distinguished Professor and the ... research team that developed the new test. The current ...
... percent increase in lifespan, raising the tantalizing question of ... taking an anti-aging medication. Results of the Buck Institute ... are currently published online in the Journal of Biological ... mood affective disorders, including bipolar disease for decades. While ...
... Monmouth University,s Urban Coast Institute will host its ... 2007, from 10 a.m. to 12 p.m., in the ... an Ocean Literate Society: Building Communities for Action," will ... of communication will be used to increase the general ...
Cached Biology News:Iowa State researchers develop technology for early detection of viruses 2Drug commonly used to treat bipolar disorder dramatically increases lifespan in worms 2Abel to receive posthumous Ocean and Coastal Leadership award 2
... Thermo's Wellwash AC is the easiest ... and is provided with liquid level ... bottles. The integrated orbital shaker and ... excellent washing performance. The big bottle ...
... Labsystems' Wellwash AC is the ... washer available, and is provided ... wash, waste and rinse bottles. ... automatic rinsing and priming functions ...
The Wellwash 4 Mk 2 microplate washer combines flexibility, simplicity and ease of use. This reliable microplate washer meets the exacting standards of today's ELISA assay requirements....
... Rabbit polyclonal to MLF1 Interacting Protein/PBIP1 ... all tested applications). ... to amino acids 73-83 of Human ... Entrez Gene ID: 359815 ...
Biology Products: