Method Development for a Local Anesthetic (A Case Study) An analyst was using a Cogent Diamond Hydride™ Column for evaluating Lidocaine methods and although they had good retention of the compound, they did not meet the required 0.3 – 0.5 AU when 0.1 mg/mL Lidocaine sample was injected for both 2μL and 10μL injection volumes. […]
Hypoglycin A and Methylenecyclopropylglycine are two compounds found in the extract of Litchi Seeds that studies have shown to be toxic. Methylenecyclopropylglycine inhibits the synthesis of fatty acids from Glucose, and therefore undernourished children are more at risk since their Glucose reserves are low. This study used LCMS with a Cogent Diamond Hydride™ Column to […]
Agilent 1290 Infinity II is stated to be a universal system that is compatible for any HPLC or UHPLC applications. The Cogent 2.o™ HPLC columns are considered “near UHPLC” with pressure limits approximately 9,000 psi. If you are using Aqueous Normal Phase (ANP) HPLC it is unlikely you will develop such high pressure. Note: The […]
If you are using LCMS, the simple answer is yes but pH is also something to consider. Acetone has a high UV cutoff and therefore can be a problem for UV detection. However, this is not a problem when using LCMS or ELS detectors such as the Corona CAD. You can use acetone to replace acetonitrile with the Cogent TYPE-C™ […]
Ion-pairing will not work with the Diamond Hydride column. It separates on the basis of polarity. Using an ion-pairing reagent reduces that polarity so the compound can be separated on a reversed-phase column.
Yes, but there is some evidence that the use of phosphoric acid may semi-permanently change the surface chemistry, and therefore the chromatography of the column. For example, if you get a certain separation using formic acid and then introduce phosphoric acid to the column, you may not get the same separation when you try the […]
Using only Acetonitrile and Water and no additives is fully compatible with the Column and will not damage it. We might however recommend having 0.1% Formic Acid during Column Storage to help inhibit bacterial growth. You should change your Aqueous Solvent more frequently in this case for the same reason. If you find the Peak […]
We are looking for a column capable of separating cocaine and its metabolites particularly Ecgonin which is very polar. The remaining metabolites (BE, EME, etc.) are less polar. The idea would be to separate all of them in one run. Any suggestion which type of Cogent TYPE-C column would be appropriate for direct injection of […]
In terms of storage, conditioning, and general use, the same protocols can be used for the Cogent Diamond Hydride 4um and Cogent Diamond Hydride 2.o™ products. The main difference between the two is of course particle size. Cogent Diamond Hydride Product Page
Reversed Phase & Aqueous Normal Phase Method Tips: You can use the Cogent Bidentate C18™ column in reversed phase (RP) with a high water content method. According to a 2014 third-party research article, if the analysis is done by reversed phase an ion pair agent is recommended to increase retention and reduce peak tailing. Here, […]
What suggestion can you offer for Analysis of Metabolites such as Glucose, Pyruvate, Taurocholic Acid, Gamma Glutamylcysteine, Glycochenodeoxycholate, Glycerol-3-phosphate, Spermidine, Taurine, and S-Adenosylmethionine? Suggestion: We would recommend starting by using the Cogent Diamond Hydride™ HPLC Column for this Analysis. Many of these compounds are quite Polar and would be very difficult to Separate by Conventional […]
How can I restore my Diamond Hydride HPLC column? I ran out of mobile phase in my B solvent and air was accidently introduced into it and my analytes are not retaining now.
Although you should never let any HPLC column dry out, your column is probably not ruined. However, It could take days to get it restored when conditioning it with your mobile phase. If there is air in the column, give it a few hours of slow flow rate of your mobile phase or use isopropanol […]
How do I Avoid Distorted Peak Shapes when using a Basic Mobile Phase pH for the Analysis of a Lipophilic Amine? – FAQ
You may not require a Basic Mobile Phase pH for HPLC analysis of these Types of Analytes. Use of the Cogent Diamond Hydride™ Column in ANP Mode with 0.1% Formic Acid instead of a Basic Mobile Phase may be a better approach. There are a number of possible reasons that you may have observed a […]
For all new Cogent TYPE-C™ HPLC columns with the exception of new Diamond Hydride™ columns, all one needs to do is run 7-10 column bed volumes of your mobile phase through the column at your normal flow rate. Then it is best to inject a known standard and repeat the injections until duplicate chromatograms are […]
How do Retention and Efficiency of Asymmetric Dimethylarginine, ADMA Compare Using 4um vs. 2.o™ Diamond Hydride™ Columns?
ADMA can be retained using an ANP gradient method with the Cogent Diamond Hydride™ HPLC column. Retention times for ADMA differed only slightly between the 4um and 2.o™ stationary phases. Efficiency however was notably higher when using the 2.o™ phase. This can be readily observed from the greater peak height using the 2.o™ column in the […]
How do retention and efficiency of glucosamine compare using 4um vs. 2.o™ Cogent Diamond Hydride™ HPLC columns?
Glucosamine can be retained using an ANP gradient method with the Cogent Diamond Hydride™ HPLC column. Using either 4um particle size or 2.o™ (2.2um) columns did not affect retention times but efficiency was notably higher when using the 2.o™ phase. This can be readily observed from the greater peak height using the 2.o™ column in the chromatogram […]
The Diamond Hydride™ column comes with an insert that describes initial conditioning steps that should be done when you first receive the column. These washings usually only need to be done once to condition the column but that will depend on what you inject and how well you prepare and filter your samples. Under normal […]
For Serum and other biological samples, we Recommend using: Solvent A: 50% DI Water/50% Methanol (or 2-propanol) + 0.1% Formic Acid (for positive mode MS) or +10 mM ammonium acetate or formate (for negative mode MS). Solvent B: The same as you would ordinarily use. Cleaning Tip: Methanol or 2-Propanol (IPA) helps to clean the […]
Before the First use of the Diamond Hydride Column: 1. Condition it with 50:50 MeOH/Water for 30 minutes. Conditioning Steps – Depends on Your Samples: 1. When using urine, plasma, or other biological samples etc: A. Conditioning the Column with 80% Acetonitrile/ 20% DI water/ 0.2% Acetic Acid (or 0.1% Formic Acid) is recommended for […]
I am not observing any peaks for succinic acid or alpha-ketoglutarate with the Cogent Diamond Hydride column in my LCMS runs.
Question: I am using LC-MS with positive ionization mode and the Diamond Hydride column. My mobile phase has 0.1% formic acid in A and B solvents. However, I see no peaks for succinic acid and alpha-ketoglutarate in the EICs. What is the problem? Answer: These compounds are both acidic and will not be amenable to either an […]
I am using a Cogent TYPE-C™column with LC-MS and solvents containing 10 mM ammonium acetate. I noticed that my source was getting white film on it. At the same time the background noise was very low. Is my column bleeding the packing material?
Cogent TYPE-C™ columns including the Cogent Diamond Hydride have a very low background noise even when used with LC-MS and the white film most likely comes from ammonium acetate build up from your mobile phase. These columns have been proven to be extremely durable and because of the direct silicon-carbon bonds of these phases, bonded phase bleed is almost […]
I was analyzing maleic and fumaric acids in biological extract using a Cogent Diamond Hydride column. After 10 perfect injections (%RSD of retention times around 0.5), I noticed that the peaks intensity diminished about 10 times, however the peaks still have the same retention time. What happened to the column?
Most likely, nothing happened to the column. Your mass spectrometer probably needs some attention. I would clean the ion source and check the nebulizer.
Question: I am analyzing S-adenosyl methionine (SAMe) with the Diamond Hydride column in ANP mode. I used a gradient with a formic acid additive and then tried an ammonium acetate additive. With ammonium acetate, I got lower retention than with formic acid using otherwise identical method conditions. I was expecting greater retention with ammonium acetate since the carboxyl […]
I have recently been using the 2.1 x 150mm 4um 100A Cogent Diamond Hydride™ HPLC Column. I am using a 400uL / minute flow rate with Acetonitrile and an Aqueous Buffer (Column Temperature = 30 °C). I’m seeing a very low back pressure (around 20 bar at the beginning of the gradient while acetonitrile level is […]
There are many differences between the many HILIC columns and Cogent TYPE-C™ columns but for the sake of brevity, I will only address the main differences. Similarity: The Cogent TYPE-C (Silica Hydride) HPLC Columns perform similarly to HILIC columns as far as polar compound elution order is concerned (when using higher than 70% organic composition of the […]
The Maximum Operating Temperature for the Cogent Diamond Hydride™ HPLC Column is 60° C. However, Higher Temperature Columns are available on request with High Temperature all Stainless Steel Hardware. Click HERE for Cogent Diamond Hydride Ordering Information
Question: I am analyzing pyrazinoic acid (see structure below) by Aqueous Normal Phase HPLC (ANP) using the Cogent Diamond Hydride™ column. I observe two broad, split peaks for the analyte. I am using an acetonitrile / DI water / formic acid based mobile phase and my diluent is 50:50 methanol / acetonitrile. Retention is also low. What can […]
Retention and/or peak shape for citric acid has changed compared to data from a previous HPLC run. What happened?
Citric acid analysis using LCMS can be compromised due to the presence of iron in the system. When iron is present, it can cause peak distortion for compounds like this. A possible solution to this problem is to use a small concentration of a chelating agent (EDTA) in the mobile phase/sample diluent to sequester the […]
The Recommended pH Range for the Cogent Diamond Hydride™ HPLC Column is 2.5-7.0. Click HERE for Cogent Diamond Hydride Column Ordering Information
Tip – Restoring the Initial Results with a Method for Analyzing AMP, ADP, ATP, UDP and GPT in Biological Extracts.
“My Results deteriorated after two days of using the Cogent Diamond Hydride Column for AMP, ADP, ATP, UDP and GPT in biological extracts”. Phosphate containing metabolites that you are using are very sensitive to the presence of sodium in your LC-MS system. After few days there is enough Sodium in your System (leaching from the […]
When using a gradient method with the Cogent Diamond Hydride™ HPLC column, if an inconsistent baseline may be observed… The following are some possible causes of the issue along with suggestions to fix it. 1. Is the mobile phase filtered before use on the instrument? A 0.45µm nylon membrane filter with vacuum filtration is recommended […]
Using the Cogent Diamond HPLC Hydride column, will Na+ interfere with succinate when analyzing sodium succinate?
No. Sodium ions, Na+ will normally elute at or near the void volume in an ANP method using the Cogent Diamond Hydride™ column. It should not interfere with quantitation of the succinate peak or any other peak for that matter. Cogent Diamond Hydride Column Ordering Information
Using Two Columns and two Modes of HPLC Together If you connect the Cogent Bidentate C18™ & Cogent Diamond Hydride™ columns in parallel, but use the same mobile phase program on both columns, it will be difficult to have both Reversed Phase and Aqueous Normal Phase (ANP) modes operating simultaneously, therefore you will not get […]
What are some examples of third-party journal articles that feature the Cogent™ HPLC columns in metabolomics studies?
There are many examples of the Cogent™ HPLC columns used in metabolomics research, which have been published by third-party researchers in peer-reviewed scientific journals. Two notable works are described here. The first one, performed by Dr. Kyu Rhee and co-workers of Weill Cornell Medical College, uses the Diamond Hydride™ column to understand the pathogenic mechanism […]
Aqueous normal phase chromatography (ANP) is a chromatographic technique which encompasses the mobile phase region between reversed-phase chromatography (RP) and organic normal phase chromatography (ONP) used mainly to separate polar compounds such as acids, bases and peptides.. Using a hydrophobic stationary phase such as silica hydride (available as Cogent TYPE-C™ HPLC columns) a mobile phase of 98% organic […]
There is less than 2% carbon load on the column. However it should be noted that the carbon on this column is not directly responsible for separation. While the carbon does modify the surface and the retention character of the columns it is not involved in hydrophobic interaction and therefore not directly responsible for retention. […]
In ANP, the strongest solvent is water. The order in decreasing solvent strength is: DI water > methanol > isopropanol > acetone ~ acetonitrile These are general estimates; in this comparison of acetone and acetonitrile for different amino acids, retention is somewhat comparable for some but significantly different for others. For methanol or isopropanol, these are typically […]
What Pressure Should I Expect with a Diamond Hydride 2.o HPLC Column at Flow Rates From 0.2-2.0 mL/min?
The below data was recorded on a Waters UPLC™ system. Please note that the pressure can vary widely depending on such factors as mobile phase viscosity, temperature, column ID, and column length. The conditions shown below should clarify any questions you may have regarding this data. Column: Cogent Diamond Hydride™, 2.2um 120A, 2.1 x 50mm, cat# […]
There can be many reasons why any compound does not retain in any given HPLC Method, however in this case, Triphenylamine has three bulky Phenyl Groups (see structure below), which can create steric hindrance from interaction of the Stationary Phase with the Tertiary Amine. NOTE: Tertiary Amines are the least Polar Amine and will have […]