Retain Highly Polar Compounds with LCMS – AppNote

Analysis of Amino-Caproic Acid by a Simple LCMS Method 6-Amino-N-Caproic Acid is an active pharmaceutical ingredient used in massive thrombolysis and proteolysis secondary to metastatic carcinoma of the prostate. It is also a potent in vitro inhibitor of fibrinolysis. After oral administration 6-Amino-N-Caproic Acid enhances the uptake of labeled Fibrinogen in both the Walker and […]

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Doxycycline Analyzed with HPLC – AppNote

Forced Degradation Method Using Shape Selectivity Doxycycline is known to epimerize at the C4 and C6 positions in acid conditions. Epimers can be difficult to separate in Reversed Phase due to subtle differences in hydrophobicity. However, this Method differentiates based on shape as well as hydrophobicity and is therefore well suited to this Isomer separation. […]

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Glipizide Analyzed with HPLC – AppNote

Separation of API from Acid Degradation Products This Method illustrates the capabilities to separate several Glipizide Degradants that are formed under acidic conditions. Good separation is obtained as well as Sharp, Symmetric Peak shapes for each compound. The Retention times show good repeatability as well, as demonstrated in the Figure overlays. Also, the Mobile Phase […]

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Cefprozil E & Z Isomers Analyzed with HPLC – AppNote

LCMS Compatible Method The USP Assay Method for Cefprozil uses a phosphate-based Mobile Phase, which is not compatible with LCMS. In this Method, only Formic Acid is needed in the Mobile Phase. The Method meets the isomer resolution criterion of not less than 2.5 with an average calculated value of 4.4. The tailing factor was […]

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Ethambutol Analyzed with LCMS- AppNote

Retention of Hydrophilic Analyte in a Drug Product When Ethambutol is analyzed on many C18 Columns, it strongly interacts with residual Silanol groups in the Stationary Phase. These interactions normally cause excessive Retention and severe Peak broadening. In this LCMS Method, Formic Acid is all that is required for the analysis of charged drugs like […]

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Celecoxib Analyzed with HPLC – AppNote

Impurities Method with Improved Peak Shape Celecoxib is Separated from an Impurity in this simple Gradient Method. The Peak shape obtained for this API shows good Peak Symmetry. In addition, the Method shows good repeatability from run to run. The API Retention time %RSD from five runs was 0.025%. Figure A shows the excellent Precision […]

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Clopidogrel Analyzed with HPLC – AppNote

LCMS Compatible Separation of API from Degradants Although Clopidogrel, commonly known as Plavix, retains well in Reversed Phase, its Degradants showed low Retention even at high water content. Therefore this Method the compounds are retained on the basis of polarity. This Method uses MS compatible Solvents and provides good Retention and separation between the API […]

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Methotrexate Analyzed with LCMS- AppNote

Methotrexate an Anti-Neoplastic and Anti-Tumor Drug The powerful anticancer drug, Methotrexate (4-Amino-N10- Methylpteroyl Glutamic Acid) acts as an antimetabolite and is used for the treatment of many neoplastic diseases including acute leukemia, osteosarcoma, non-Hodgkins lymphoma, and breast cancer. There is a great interest in pharmacological studies and clinical monitoring of Methotrexate. A quadrupole Mass Spectrometer […]

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Domoic Acid in Seafood Samples Analyzed by LCMS – AppNote

Domoic Acid (DA) From Seafood Without Derivatization Several Seafood and Plankton samples were tested for Domoic Acid (DA) which, as a highly polar compound, is poorly Retained on most Reversed Phase Columns. This developed Method based on HPLC and MS detection does not require derivatization of Domoic Acid. The concentration of this potent toxin varied […]

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Fexofenadine Analyzed with HPLC – AppNote

Simple Isocratic Assay Method without Amine Additives Fexofenadine, antihistamine, has a number of aromatic moieties and the tertiary amine in its chemical structure that can cause Peak tailing issues when using most L11 Columns for Analysis. The USP Assay Monograph for Fexofenadine Tablets uses Triethylamine to reduce tailing. With this Method, Trifluoroacetic Acid is used […]

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Aspartic Acid & Arginine Analyzed with LCMS- AppNote

Reduced Sample Prep and Handling Two amino acids are adequately Retained and can be easily quantified in this Method with very little Sample Prep. There is no need for any Pre-Column or Post Column derivatizing due to Mass Spectrometric Detection can detect these molecules and Aqueous Normal Phase (ANP) HPLC can Retain in their ionized […]

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Citric, Aconitic & Maleic Acids Analyzed with LCMS – AppNote

Low Molecular Weight Organic Acids Low mass Acids were retained and Separated in this Method with the Aqueous Normal Phase mode of HPLC. The Method was able to resolve the three compounds in 10 minutes. Limits of Detection (LOD) for them ranged from 0.05 to 24 micromolar. Peaks: 1. Maleic Acid 115.0031 m/z (M-H)- 2. […]

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Phenylglycine & Phenylalanine Analyzed with HPLC – AppNote

Analysis using a Simple Mobile Phase. In this Method two important amino acids: L-(+)–alpha-phenylglycine and L-phenylalanine, were Separated. C18 Columns used today which may be present in every Analytical Laboratory may not be able to Retain underivatized Amino Acids. They usually elute at or near the “void volume” with other polar compounds. Peaks: 1. L-(+)-Alpha-Phenylglycine […]

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Melamine Analyzed with LCMS – AppNote

Detect Trace Levels of Melamine The unique chemical properties of Melamine (base functionalities) present challenges in obtaining Retention and adequate Sensitivity, when using standard HPLC Columns. In this Method, Melamine is Analyzed under Aqueous Normal Phase (ANP) conditions with an increased Sensitivity making it possible for detection at Trace Levels. Peak: Melamine m/z 127 (M+H)+ […]

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Gentian Violet Analyzed with HPLC – AppNote

Separation of Related Compounds Gentian Violet degrades in solution, forming complex set of Degradation products. This Method can easily separate out the various compounds formed in Solution. The figure below shows a USP Standard of Gentian Violet after stressing it in solution thus creating Degradation products. With excellent Selectivity well as great Peak shape this […]

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Dipyridamole Analyzed with HPLC – AppNote

Several Tertiary Amines on a Molecule that is Easily Retained This Method for the Analysis of Dipyridamole USP Tablet Formulation is easy to do. Note the excellent Resolution from capsule excipients, along with the excellent Peak shape for this complex, difficult molecule.   Peaks: 1. Dipyridamole 2. Tablet Excipients Method Conditions Column: Cogent Bidentate C18™, 4μm, […]

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Simvastatin Analyzed with HPLC – AppNote

Excellent Peak Shape and Resolution This Method is easy to prepare, use and reproduce. Good Peak shape and Resolution is observed for Simvastatin from its hydrolysis products and tablet Excipients. Peaks: 1. Excipient 2. Hydrolysis Product 3. Simvastatin Method Conditions Column: Cogent Bidentate C18™, 4μm, 100Å Catalog No.: 40018-75P Dimensions: 4.6 x 75mm Mobile Phase: 52% DI Water / […]

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Aspirin & Salicylic Acid Analyzed with HPLC- AppNote

Excellent Peak Shape For a Very Difficult Compound This Method is easy to prepare, use, reproduce and a good Separation of Aspirin from its major hydrolysis product, Salicylic Acid is achieved. Peaks: 1. System Peak 2. Aspirin (Acetylsalicylic Acid) 3. Salicylic Acid Method Conditions Column: Cogent Bidentate C18™, 4μm, 100Å Catalog No.: 40018-75P Dimensions: 4.6 x […]

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Succinic Acid Analyzed with LCMS – AppNote

Isocratic or Gradient The analysis of Succinic Acid is mostly performed by Gas Chromatography (GC), Enzymatic Assays, Ion-Exclusion Chromatography (IEC) or by Reversed Phase HPLC. However, these techniques are time consuming and analysis time is very often longer than 1 hour. In this Application Note, a rapid, cost effective screening Method for the detection of […]

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Isobaric Metabolites in Urine Analyzed with LCMS – AppNote

A Simple and Reproducible LCMS Method This Method demonstrates the ability to separate a wide variety of polar metabolites using Mass Spectrometry for detection. When monitoring the EIC at 133 m/z, seven Peaks were positively detected in a Urine sample by comparison with standards or by using accurate mass. Although primarily focused on the analysis […]

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Chlormequat & Mepiquat Analyzed with LCMS – AppNote

Plant Growth Regulators CQ & MQ Retained and Separated Analysis of Chlormequat and Mepiquat can be challenging since they are extremely hydrophilic and are only weakly Retained in Reversed Phase HPLC which makes their Separation difficult. In addition, these compounds are Retained too strongly in Normal Phase (Non Polar Solvents) Chromatography. Due to this situation, […]

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Organic Acids Analyzed with LCMS – AppNote

An Important Class of Compounds Organic Acids, an important class of Metabolites, are very well Retained and Separated in this Method. Figure A shows the combined EICs for five small Organic Acids: Maleic, Aconitic, Fumaric, Citric and Oxaloacetic Acids. Two of these acids are isobaric compounds; Maleic and Fumaric Acids (115 m/z) and are separated […]

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Glyphosate Analyzed by LCMS – AppNote

Retention of an Extremely Polar Compound A reliable Method for the determination of Glyphosate is presented. This Method provides very reproducible Retention and fast equilibration even when a Gradient analysis is used. The use of LCMS detection avoids time-consuming derivatization of this compound which is lacking a chromophore for UV detection. The Method shown, with […]

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Peptide Mixture Separated with HPLC – AppNote

Precise and Fast Equilibration Time HPLC is a technique used for the characterization of Peptides and Reversed Phase HPLC is employed for the initial analysis and final, large scale purification. The first step of the production of Synthetic Peptides usually involves an initial Separation of these compounds from its mixture, on an analytical scale, then […]

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Cytochrome C Analyzed with HPLC – AppNote

From Horse and Bovine Heart Using the simple Reversed Phase HPLC Gradient Method shown in this note, it was possible to separate Horse and Bovine heart Cytochrome C. The Peaks were well separated and symmetrical. A linear detector response was observed over 2 orders of magnitude. Peaks: 1. Cytochrome C from Horse heart 2. Cytochrome […]

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Glutamine and Lysine Analyzed by LCMS – AppNote

Two Isobaric Amino Acid as Biomarkers Found in Saliva The Chromatogram below shows an interesting aspect of analyzing real patient samples for a pair of metabolites: Glutamine and Lysine. Figure A represents 7 samples taken from patients with cancer; Figure C shows injections of samples from 7 patients with pancreatitis while Figure B represents 7 […]

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Folic Acid Analyzed by LCMS – AppNote

Minimized Sample Prep & Increased Sensitivity for Vitamin B9 A sensitive and reliable Method was developed for the determination of Folic Acid (FA) and can be used in many applications including complex sample matrices without the Solid-Phase Extraction clean up. Since the method uses LC-MS (negative-ion) it provides sensitivity as well as great selectivity. The […]

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Paracetamol, Aspirin and Salicylic Acid HPLC Method – AppNote

Analgesic Drugs Analyzed with HPLC A robust and rapid method was developed for the accurate simultaneous determination of Paracetamol, Aspirin, Caffeine, Benzoic Acid as an internal standard and Salicylic Acid (hydrolysis product of Aspirin). This method is rapid and sensitive and therefore suitable for routine quality control of these drugs in dosage forms and for […]

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DMAP, 4-Dimethylaminopyridine Analyzed with HPLC – AppNote

Analyzing the Catalyst, DMAP, 4-Dimethylaminopyridine with HPLC This AppNote shows a rapid, reliable, Isocratic Reversed Phase, HPLC Method that was developed for the Analysis of an important Catalyst for many applications and industries: 4-Dimethylaminopyridine (DMAP). Peak: DMAP Method Conditions Column: Cogent Diamond Hydride™, 4μm, 100Å Catalog No.: 70000-7.5P Dimensions: 4.6 x 75mm Mobile Phase: —90:10 Solvent A / […]

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Efficiency of ANP Measured – AppNote

Aqueous Normal Phase (ANP) Efficiency Measured with Sorbitol, Glucose & Uracil Figure A is an excerpt from a study that investigated analyses of Glucose and Sorbitol on TYPE-C Silica based HPLC Columns and found that the efficiency of ANP is better than observed with HILIC. A Cogent Diamond Hydride Column (2.1 x 150mm with a […]

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Urea Analyzed with HPLC- AppNote

Fast, Reproducible Method In this Method Urea was retained when a 100% DI Water Mobile Phase was used. DI Water can be used as the complete Mobile Phase without fear of phase “de-wetting” and subsequent loss of retention time. This Method has the advantage of being very simple, without time consuming derivatization or reaction steps […]

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Effect of Sodium Acetate for Glucose Ionization in LCMS – AppNote

Improved Signal to Noise for Glucose with LCMS Glucose, a simple monosaccharide, was analyzed by LCMS and the peak is very symmetrical and easy to integrate. This application note illustrates the importance in the addition of Micro Molar amount of Sodium to the Mobile Phase when sugars are analyzed. Sodium adducts of sugars produce much […]

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Maleic & Fumaric Isobaric Acids Analyzed with LCMS – AppNote

Separation & Detection of Isobaric Compounds Two Isobaric Acids, Maleic and Fumaric (115 m/z) are separated when using the solvents below and a Gradient Method. Solvent A used in this application note contains 50% Methanol which is recommended when biological samples are used. This Method is reproducible and is fast to equilibrate between gradient runs […]

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Atropine Analyzed with a Gradient Method – AppNote

No Ion Pair Reagent Necessary Chromatographic separation and quantification methods of Atropa Alkaloids are often described in the literature and the method of choice is usually Ion-Pair Chromatography (IPC), which requires long equilibration times and it is not very robust. This method shows a symmetrical peak for Atropine using a simple Gradient Method that does […]

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Meprobamate Analysis by LCMS – AppNote

Rapid & Simple Analysis of Meprobamate. One Step Sample Preparation with Plasma or Urine Several methods have been reported in the literature for the quantification of Meprobamate in biological samples (plasma, urine, hair etc) for patient monitoring and other tests. Most of these methods present the drawbacks of long analysis time and complex sample pre-treatment […]

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Determination of Atropine with HPLC – AppNote

Rugged & Fast Without Paired Ion Reagents Ion-pair Chromatography (IPC) is commonly used in order to retain Atropine on ordinary Reversed Phase HPLC Columns. Beside long equilibration times with these columns, IPC often suffers from poor robustness. Using an isocratic elution with flow rate 1mL / minutes gave symmetrical peak for this Tropane Alkaloid. The […]

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Folic Acid Content in Fortified Cereals Analysis with HPLC – AppNote

Online Cleanup of Sample or Matrix Components This application note demonstrates an effective means of separating and resolving Folic Acid from undesired matrix components in cereal extracts without the use of SPE or other sample cleanup techniques. SPE is normally performed to remove compounds from the matrix which would co-elute with Folic Acid before injection […]

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Gabapentin Drug Product Analysis by LCMS – AppNote

Isocratic Retention with LCMS Gabapentin was retained successfully with a very symmetrical peak. Two different Mobile Phases were used and the advantage of the Mobile Phase used in Figure A is shown to be that Isopropanol present in the solvent A helps clean the Column which can be useful if biological samples containing lipids are […]

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Carisoprodol & Meprobamate Analysis by LCMS – AppNote

Analysis of Closely Related Compounds It has been reported that for the quantitation of Meprobamate and Carisoprodol, (in biological samples plasma, urine, hair etc) by LCMS, excessive analysis time and complex sample treatment (derivatization, solid or liquid phase extraction) may be required. The Method in this AppNote shows a rapid way to quantitate both drugs […]

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Forced Degradation of Atorvastatin by LCMS – AppNote

Separation of API from its Lactone Degradation Product Atorvastatin is separated from its main Degradation product in this Method with a simple linear Reversed Phase Gradient. With the use of LCMS, the identity of the Degradant can be confirmed from its m/z value. The Degradation is an intramolecular Fischer esterification, which is catalyzed under Acidic […]

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Fluoxetine Orthogonal Assay Method – AppNote

Simple Methods Without the Use of Ion-Pairing Agents Unlike the USP assay method, Ion Pair Agents are not needed in this application. Both figures show how good Peak symmetry can be achieved either in the Reversed Phase (RP, Figure A) or Aqueous Normal Phase (ANP, Figure B) mode with only Formic Acid as the Mobile […]

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Levothyroxine Analysis – AppNote

Superior Resolution, Reproducibility, & Peak Shapes Compared to USP Method The USP assay method for Levothyroxine requires that a resolution of not less than 5.0 must be demonstrated between Levothyroxine and related compound Liothyronine. A chromatogram obtained from following the USP method using a Type-B Silica based L10 Column is shown in Figure B. The […]

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Sarcosine Analyzed with LCMS – AppNote

Separation of Potential Urine Biomarker from Isobaric ß-Alanine This developed LCMS method can separate Sarcosine from Beta- Alanine in serum and urine samples without using labor intensive sample derivatization. Since Sarcosine is considered a potential biomarker for prostate cancer risk and aggressiveness, it is essential to resolve and accurately quantify this compound in the presence […]

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Domoic Acid Analyzed by LCMS – AppNote

Determination without Derivatization To date, several analytical methods have been developed for the quantitative determination of Domoic Acid (DA). However, many of the methods require tedious and labor intensive derivatization of this compound in order to detect it. A new method based on HPLC and MS detection which does not require derivatization has been developed. […]

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Proline: Trans-4-Hydroxy-L-Proline by LCMS – AppNote

Rapid, High Efficiency Method by LCMS This method is highly specific, efficient and fast for the analysis detection, small volumes of the reaction mixture can be injected and of trans-4-hydroxy-L-proline. Due to the high specificity of the the amount of the produced compound can be determined. No derivatization is required for the detection of this […]

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Propantheline Bromide HPLC Methods- AppNote

Orthogonal Methods without Ion-Pairing Agents In the USP assay method for Propantheline Bromide tablets, Sodium Dodecyl Sulfate (SDS) is used as an Ion-Pairing Agent in the Mobile Phase. The long alkyl chain of this additive makes it particularly difficult and time consuming to load and remove from the HPLC column. Is this method, not only […]

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Shikimic Acid Analysis by LCMS- AppNote

Ingredient for Production of Oseltamivir The method shown in this application note assures both high specificity and high sensitivity for Shikimic Acid, a synthetic ingredient for the production of the pharmaceutical Oseltamivir marketed as Tamiflu®. Using this LCMS friendly method allows for a 10-100 fold increase in sensitivity over some other methods. Also note that […]

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Hormone Replacement Capsule Analysis with HPLC – AppNote

Separation of Estriol, Estradiol, & Progesterone This gradient method features a separation of the three components of a hormone replacement formulation. Figure A shows a five run overlay of the formulation extract injections. Figure B shows a zoomed-in view so that the Estriol and Estradiol Peaks, which are present in much lower concentration than Progesterone, can […]

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Cough Syrup Ingredient Analysis with HPLC – AppNote

Separation of Antitussives, Analgesics, Decongestants, and Preservatives Cold and cough formulations may contain a number of components such as antitussives (Dextromethorphan), decongestants (Pseudoephedrine, Guaifenesin), analgesics (Acetaminophen), and preservatives (Methyl Paraben, Propyl Paraben, Benzoic Acid). The method illustrates good separation between a variety of these compounds, also that symmetric Peak shapes can be obtained in each […]

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