Hamilton Syringes Clogged on 6890N GC

After finishing the assay of methylbenzene residue in Taxol (Paclitaxel), found the 10 uL syringe clogged.
The injection concentration is 0.2 g/ml. Maybe the high injection concentration is the reason why the syringe clogged.

Option 1: Try to put the syringe into MeOH with ultrasonic bath. But failed, the MeOH had difficulty to penetrate and dissolve the blockade.

Option 2: Put the needle in DMF bath for 2 hours, then if doesn’t work well, put ithe boiling water ( Boiling IPA) for 30 min. It worked.

Option 3: (learned it from Hamilton Tech Service) Put the clogged needle into MeOH or EtOH with ultrasonic bath over night.

Today, got a calling from a friend @ A****

He commented that my Sample Pre-treatment by Solid-Phase Extraction is GOOD, but complained for a average of 15 $ on the SPE per sample.

Ok, here is another alternative for Sample Pre-treatment, I named it as dispersive solid-phase extraction.

(1 ) Extraction: Transfer 15 g of ample, accurately weighted, in 50-mL PS Centrifuge tube. Add 15-mL Glacial Acetic Acid-Acetonitrile (1:1), 6 g of anhydrous Mg2SO4, 1.5 anhydrous Na2SO4, then shake it mechanical 1 min, centrifuge the mixture for 5 min under 5000 r/min for further purify.

(2) Purification: Weight 0.1 g PSA，0.1 g C18，0.3 g of anhydrous Mg2SO4 into a 5-mL glass centrifuge tube, then pipet 2-mL of clear solution into the tube.Mixing the solution by using multi-function mixer whirlpool for 1 min, centrifuge the mixture for 1 min under 5000 r/min . Pipet 1-mL of the upper clear solution into 1 ml volumetric flask for the assay preparation.

To ensure the precision of the assay results, highly recommend the follow steps.
(3) Add the internal standard and protective agents: Put the 1 ml volumetric flask onto a Nitrogen blowing Instrument, blow with N2, concentrate the solution volume is less than 0.8-mL at room temp.Transfer an accurately weighted quantity 100 100 µL internal standard of TPP and 100-µL of protective agent’s mixture solution. Bring to the volume with Acetonitrile. Homogenize the solution on a vortex 5 min for GC-MS analysis.

Would you please update me if it works. Thanks.
Appreciate your comments.

Note:Working 200% well with the vegetable juices and fruit juices. For the Grains, Beans, Nuts, Seeds, please add more water and increase extraction time with vigorous mechanically shake, it also works.

Extraction Melamine Methods from Pet Foods (diluted 100 times) Comment (Just my 2 cents)

There are at least up to 3 prevailing assay methods for Melamine.
(1) Sublimatography
(2) HPLC Method
(3) HPLC-MS（FDA，SGS) and GC-MS(FDA)Sample Preparation:
1. Homogenize the pet foods into the shape of structural pudding.
2. Weigh 2 g of sample into 10mL MeOH/H2O( 60:40)， shake it vigorously.
3. Sonicate 1 min.
4. Mixing the solution by using multi-function mixer whirlpool, let it still 5 min, allow aqueous phase and organic phase separate.
5. Centrifuge it 5 min under 3000 g.
6. Filter the solution with whatman glass fiber filter paper and collect the filtrate.
7. Pipet 0.1 mL of filtrate and dilute with 1.9 MeOH/H2O( 60:40) and 20 mM PBS.
8. Pipet 100 uL for further GC-MS analysis.

Results: Using this sample pre-treatment method assay 8 commercial cat foods ( 15 ppm melamine label claimed, No-FDA recall Porducts), the recovery ratio is from 75%-117%.

http://www.fda.gov/oc/opacom/hottopics/petfood.html

Why the sample pre-treatment matter? the answer is obviously for those chemist whose had nightmare with extraction Melamine from aqueous phase. VIP tips: when obtained a poor recovery result, DO NOT blame the GC-MS, pay attention to the that Melamine solubility in H2O vary under different temperatures.

Note: this method is for products screening only, NOT for any other purpose.

Sample Pre-treatment by Solid-Phase Eextraction

Pesticide residue analysis in Grains, Beans, Nuts, Seeds
(my own method I)

Sample Pre-treatment
(1) Place about 10 g of sample, accurately weighted, in 100-mL volumetric flask, add 20 mL of D.I H2O, let it still 1 min.
(2) Homogenize 20 mL of Acetontrile with frequent shaking, then filter. Wash the residues on the filter paper by 20 mL of Acetontrile. Combine all the filtrate, dilute to volume with Acetonitrile.
(3) Piper 20 mL of filtrate into 250 mL flask, mix 10 g of NaCl and 20 mL of 0.5 mol/L Phosphoric Acid Buffer. Allow the solution cool, still, discarding the aqueous phase.
SPE Column Solvation/Pre-equilibration
(A) Activation: Activate the Bond Elut C18( 1 g ) by washing 10 mL of Acetonitrile.
(B) Load Sample: Pipet 20 mL of the filtrate, collect the elute.
(3) Rinse / Analyte Elution: Use 2 mL of Acetonitrile.
(4) Concentrate: under 40 degree C, use rotary Evaporator. Add 10 mL of a mixture solution of Acetonitrile/Methylbenzene (3:1) for further pre-treatment.
Column Purification by Carbon/NH2 SPE
(1) Column solvation: Activate the Bond Elut Carbon/NH2 (500mg/500mg) by 10 mL of a mixture solution of Acetonitrile/Methylbenzene(3:1).
(2) Apply sample: Pipet 2 mL of the above solution.
(3) Interference elution: Analyte elution by using 20 mL of Acetonitrile/Methylbenzene (3:1), collect the elute.
(4) Concentrate: All 5 mL of Acetone to re-dissolve eluates again. Under 40 degree C, evaporate on a stream bath to dryness (or with Nitrogen blowing Instrument) and dissolve the residue in 1 mL with Acetone/Hexane (1:1).
Now ready for GC-MS or GC-MS/MS.
To my best knowledge, my own method meets assay requirement for hundreds of pesticides, antiseptic, herbicides, feed additives, Veterinary Drugs that required by Japanese Ministry of Health, Labor "food maximum residue limits of pesticides in the interim standards".

Welcome email if have some technical issue....
Just for convenience, put SPE at here, not for Advertising.
Bond Elut C18 (1g, 6mL, PN.12256001)
Bond Elut Carbon/NH2 (500mg/500mg, 6mL, PN.12252202 )

Assay the Chlorothalonil & Dicofol by GC-MS with Solid Phase Extraction

Sample Preparation: Supelco C18 Solid phase Extraction.
Std Prep: pipet 25.0, 50.0, 100.0, 150.0, 200.0 ul from EPA stock STD solution ( 10 ug/ ml) into 5 ml VF, then bring to volume with acetone to obtain a serial of Conc. 50.0, 100.0, 200.0, 300.0, 400.0 ug/L STD solution.
Column: DB - 5MS ( 30 m × 0.25 mm id × 0125μm)
Initial Temp: 120℃ hold 2 min, ramp to 240℃ at 15℃/min, Hold 5 min, ramp to 260℃ at 10℃/min, hold 2 min
Carrier gas: He
Flow rate: 1 ml/min
Injection Volume: 1 ul
Split-Splitless: Splitless at 1 min, Splitl ratio: 1∶20
Mass: EI ion sources ( 70 v) ;Ion Sources Temp: 240℃;
Connect Temp: 260℃;
Solvent delay: 310 min;
Full Scan Range M/Z:40 ～ 450 amu。
Selecting Ion Monitor (SIM) for quantitative analysis ( Please see the table)
Chlorothalonil: 139, 141, 250, Molecular Ion: 139
Dicofol : 264, 266, 268, Molecular Ion: 266

http://en.wikipedia.org/wiki/Chlorothalonil

Pesticides Molecular Ion ( M/Z) Selecting Ion Monitor ( M/Z) Retention Time
Chlorothalonil 268 264, 266, 268 10.5
Quintozene 249 214, 247, 249 8.5
Dicofol (DDT) 139 141, 250 139 14.2

Assay the Methylbenzene Residue in the Taxol (Paclitaxel)

GC: Agilent 6890N- FID
Column: J&W DB-624 GC Column 75m, 0.53mm, 3.00um.
Oven Temp Ramp:
Initial Temp 35 degree C and hold 6 min, ramp to 150 at 40 C/min, hold for 10 min.
Constant Flow rate: 5mL/min
Inlet Temp: 200 degree C
FID Temp: 250 degree C
Solvent: Dimethylfomamide (DMF)
Injection Volume: 1 ul
Split Ratio:1:50
Carrier gas: N2（99.999%）

Found the assay information from the LabNote#53027 in Tech 2005, so missing the time @ Tech: Work Hard, Play Crazy, Study Smart..

Identification Test: Goldenrod

Goldenrod

Identification Test: Transfer 1 g of sample in 20 ml of Petroleum ether and sonicate 20 min to a suitable flask, filter the sample to remove the Petroleum ether. Allow it dry, add 1 ml diluted HCl and 50 ml Ethyl Acetate, then sonicate 30 min. Filter the sample to remove the solvent, allow the residue to dry, add 2 ml MeOH, the residue dissolve.

Apply separate 10-ul portions of this solution and of a standard solution of USP Chlorogenic Acid RS in MeOH containing 0.5 mg per ml to the starting line of a thin-layer chromatographic plate (See chromatography <621>) coating with 0.25-mm layer of chromatographic silica gel mixture. Allow the spots dry, and develop the chromatogram in an unsaturated chamber with a solvent system consisting of a mixture of Toluene-Ethyl Acetate-Formic Acid-Glacial Acetic Acid-H2O is 1/ 15/ 1/ 1/ 2) until the solvent front have moved about 3/4 of the length of the plate. Remove the plate from chamber, air-dry and view under 365 nm long-wavelength UV-Light: the Rf value, color of the principal spot obtained from the test solution corresponds to that obtained from the standard solution.

From China Pharmacopoeia 2005 Volume 1- P218 .

If need,
HPLC Analysis of the flavonoids in pharmaceutical preparations from Solidago canadensis
See the paper: J. Pharm. Biomed. Anal. 32 (2003) 1045-2-53.
Results: NLT 0.20 % Chlorogenic acid in goldenrod....

Done....Night--- 11:05 PM July-15-2008

Linear-Solvent Gradient Dlution in HPLC

The linear-solvent-strength (LSS) model for gradient elution is based on an approximation for isocratic retention in RP-LC as a function of solvent strength.

Bonus Question: When run out of mobile phase, what is the consequence for the HPLC system...
A: all the hydraulic pathway would dry out
B: the air come into the pump, hydraulic pathway
C: the air would come into Column
D: the air could come into flow cell.
or what else....
( Tip: High school physical knowledge: air-pump...or try to recall when onsite setup the HPLC system, what need to do b4 start the pump at the first time?
Cool, one coke for the right answer, Mountain Dew

Peak Purity Analysis by Chemstation (2)

Peak Purity Analysis by Chemstation (1)

A key questions in general for chromatography and in particular for drug discovery is whether a given peak of interest represents co-eluting components. By using an algorithm in the LC/MS ChemStation to determining the number of components in a chromatographic peak. ( Just calculate a numerical value to characterize the degree of dissimilarity of the peak spectra, a so-called similarity factor, based on the match of the peak spectra to one another.)Two Curves

(1) Similarity curve: The mathematical fundamentals used in the similarity curve calculations are those used for the purity factor, however they are displayed in another format. All spectra from a peak are compared with one or more spectra selected by the operator, an apex spectrum for example. The degree of match or spectral similarity is plotted over time during elution. An ideal profile of a pure peak is a flat line at 1000.

(2) A threshold curve: shows the effect of noise on a given similarity curve. The effect increases rapidly towards both ends of the peak. In essence, a threshold curve is a similarity curve with background noise contribution.

In Summary, the threshold curve, represented by the broken line, gives the range for which spectral impurity lies within the noise limit. Above this threshold, spectral impurity exceeds the spectral background noise and the similarity curve intersects the threshold curve indicating an impurity providing the reference and noise parameters have been sensibly chosen.

Heavy Metals Assay: Can Not Print in my Varian 240 FS Atomic Absorption spectrometers

Today, Co-worker came to me and complained that issue of "Can not Print" " Loss the raw data" in the Varian AAS 24o FS.

Yes, there are some bugs in the Varian SpectrAA software 5.1 Pro.

How to fix : Called the technical support several times, was asked to pay attention on the report setting when "edit sequence parameters"/ then go to "Reports", select the "Print Report" and save the raw data.

That maybe help eliminate that problem.
After that, the "raw date missing" and "failed to print" gone..

Cool...