Ligusticum chuanxiong Hort

Identification:

(1) Weigh 1 g of sample in 5ml of Petroleum ether ( 30-60 degree C) into 10 ml tube, shake the tube 2 or 3 times in 10 hrs. Let it still, pipet the upper clear solution 1 ml in to a evaporating dish. Allow the solvent to evaporate, add 1 ml MeOH, the residues dissolve.

(2) then add 2% 3,5-Dinitrobenzoic Acid (in MeOH) testing solution 2-3 drops, and 2 drops of Sodium Hydroxide Solution (saturated dissolves in Methanol), display Purple & red.

From China Pharmacopoeia 2005 - P28.

Mass Spectra of Ferulic Acid (CAS# 537-98-4)
C10H10O4
Source Temperature: 150 °C,
Sample Temperature: 140 °C,
DIRECT, 75 eV,
Mass of molecular ion: 194.

Peak Data:

    51.0       5.5
   77.0       7.0
  105.0       5.7
  133.0      13.6
  145.0       5.0
  161.0       7.4
  176.0       5.2
  177.0       8.6
  179.0      19.0
  193.0       6.1
  194.0     100.0
  195.0      11.1
Low down the flow rate and injection concentration.

Dandelion

Assay of Caffeic Acid in Dandelion

Dandelion (Herb Taraxaci)

Identification: TLC.

Column : C18, 250 x 4.6, 5 um
Mobile Phase A: 0.01 M NaH2PO4, pH 3.8-4.0.
B : MeOH. A/B is 70/30.
Flow Rate: 1.0 ml/min
Detector Wavelength: 323 nm, 30 degree C.

Standard Prep: 7.5 mg std in 50 ml MeOH, shake & dissolve well, pipet 2 ml to 1o ml volumetric flask, bring to volume with MeOH.
Sample Prep: Weigh 1 g in 50 ml, fill to volume with 5% HOAc in MeOH.

Results: HPLC assay of Caffeic acid NLT 0.020 %. Correct me if i am wrong, to my best knowledge, there is some connection between Caffeic acid and carcinogenicity.

Pesticides on the Cucumber/Strawberry/Carrot

Today, just like to do something to kill the time....

The Difficulty is How to isolate/remove the polar pigments (chlorophyll and carotinoids) /fatty acids, organic acids before LC-MS/ GC-MS?

Strongly recommend the combine of PSA and GCB (PSA= primary-secondary amine, GCB = graphitized carbon black)

Steps:

(1) Sample preparation and extraction

Sample: 10g of cucumber/strawberries were homogenized and placed in a 50mL PTFE centrifuge tube.

Solvent: 10mL of acetonitrile were added to homogenate Shake for 1 minute, until uniform

Salts: 4.0g MgSO4 (anhydrous powder or granular), 1.0g NaCl, 1.0g trisodium citrate dihydrate
0.5g disodium hydrogencitrate sesquihydrate

Salts were added and vigorously shaken for 1 minute. Sample was centrifuged and the supernatant removed for cleanup. Pesticides standards (200ng/mL) were spiked in at this point

(2) Sample extract cleanup ( Here, Do not recommend any brand to hurt "Fair Play Spirit" )

1mL of supernatant from the previous step was placed into several 2mL polypropylene centrifuge tubes, each containing one of the following adsorbent mixes:

• 500 mg PSA
• 250-300 mg GCB,
• 1.2 g MgSO4
• 4.0 mL Acetone/Toluene(3/1)
• Shake and Centrifuge

Samples were shaken with the adsorbents for 30 seconds (carbon for 2 minutes), then centrifuged to produce a clear supernatant for GC/MS analysis.

GC-MS:
Column: A 30m, 0.18mm ID, 0.14µm.

Sample: Custom pesticide mix 200µg/mL each pesticide,

Inj.: 1.0µL splitless (Hold 1 min.)

Inj. temp.: 250°C

Carrier gas: Helium

Flow rate: Constant linear velocity @ 40cm/sec

Oven temp.: 40°C (hold 1 min.) to 320°C @ 12°C/min.

Det: Aiglent 6890N-5973 MSD

Transfer line temp.: 300°C

Ionization: Electron ionization

Mode: Selected ion monitoring (SIM)

Results:
Should soak/wash the fruits before eating
email me if want the assay results (not public, not panic).

Electrospray Summary

1. Analyte type:

1.1 Preformed ions (acids and bases)
1.2 Polar neutrals
1.3 Multiply charged ions of biopolymers
1.4 NMT<100>

2. Typical flow rates: low down - 1.0 ml/min
3. Promote ionization:
3.1 Correct pH
3.2 Favorable HPLC solvent composition
3.3 Post-column addition of reagents

4. Soft ionization technique

5. Typical applications: Drugs, Sugars, Peptides, Proteins, Oligonucleotides

Here is another question: Why GC-MS have a NIST/EPA/NIH Mass Spectral Library 2005 or 2008.? that is "Standard" Mass Spectral for GC-MS, it help newbie chemist would master the elucidation skill of Mass Spectral in 3 weeks.

However, Why LC-MS do not have any LC-MS Mass Spectral Library?
Answer: give me one Coke, i will tell you..... or call the Agilent/Waters/Varian/AB/PE Technical Support, they would answer...

Tips: Look at above # 4: comparing the ionization technique.
for LC-MS: ESI, APCI, APPI ( they are Soft technique), the ionization energy@interface different, the mass fragment different, ---- so not a universal mass library. Or or call the Agilent/Waters/Varian/AB Technical Support, ask them why develop so many "New and Individual" Patent ( on their own instrumentation only) on LC-MS ionization technique...

For GC-MS: EI ( Hard ionization technique), the Ionization energy (from 5 to 241.5 eV) is stable, so the molecular fragment should be the same....that is the reason why NIST/EPA/NIH have Mass Library.

It is so simple: understand the basic ionization technique, should answer.

Maximizing High Flow ESI Sensitivity

Today, met a general question: how to improve the ESI Sensitivity? So I would like to answer this question from the preparation of sample.

(1) Select appropriate chromatography grade HPLC solvents
(2) Avoid exotic solvent mixes (MeOH, MeCN, Water, 0.1% formic work best for 98% of LC/MS applications)
(3) Avoid adding excessive modifiers (eg amm.acetate @ 10mM not 50mM)
Choose the right column chemistry (C-8 vs. C-18); change column chemistry before changing solvent mix or composition.
(4) 2.1mm column or lower, flow rates of 200-400 uL/min
(5) Peak widths for quantification not greater than 8-10 secs
(6) Dissolve sample in start mobile phase solvent (weakest solvent possible).

Here is a schematic drawing of the Finnigan MAT 900S electrospray ion source (kindly provided by EPA/Dr. Helmut Muenster.)

Suitable Solvents for LC/MS

(1) Reverse-Phase LC/MS Solvents
ACN, MeOH, H2O, Isopropanol.

(2) Normal-Phase LC/MS Solvents (for APCI-MS)
Hexane, Methylene Chloride, Acetone, Ethanol.

(3) Compatible LC/MS Buffers and Modifiers:
Formic acid, Acetic Acid, Ammonium Acetate, Ammonium Formate,
Ammonium Hydroxide, Trifluoroacetic Acid (TFA) concentration should
be <0.1%.

Avoid Non-volatile Buffers, Alkali-metal phosphates, borates, etc. that is to say,

ØAcids

l Do not use inorganic acids (may cause source corrosion), Formic and acetic acid are recommended.

Ø Bases:l Do not use alkali metal bases (may cause source corrosion), Ammonium
hydroxide and ammonia solutions are recommended.

Surfactants (surface active agents): lDetergents and other surface active agents may suppress ionization.

Ø
Trifluoroacetic Acid (TFA): l May enhance chromatographic resolution, but causes ion suppression in both negative and positive ion modes.

Ø Triethylamine/Trimethylamine (TEA/TMA): May enhance deprotonation for Negative Ion Formation.

Steps for ESI Optimization (Electrospray Ionizatio)

Steps for ESI Optimization

1 If analyte’s pKa is unknown, evaluate 3 pH regions in positive and negative ion modes.

2 Acids – Negative Ion detection, adjust pH 2 units above pKa: Increase pH with NH4OH, TEA, TMA.

3 Bases – Postive Ion Detection, adjust pH 2 units below pKa* Decrease pH use formic acid , acetic acid, TFA. *In complex molecules, many exceptions to these rules are observed.

4 Remove salts which may cause ion suppression.

5 Adjust source temperature and source voltages to maximize signal

6 In negative ion mode, use lower spray voltage to minimize discharge.

The analyte is introduced to the source in solution either from the eluent flow from liquid chromatography. (Flow rate is 1µl min^-1) . The analyte solution flow passes through the electrospray needle that has a high potential difference (with respect to the counter electrode) applied to it (typically in the range from 2.5 to 4 kV). This forces the spraying of charged droplets from the needle with a surface charge of the same polarity to the charge on the needle. The droplets are repelled from the needle towards the source sampling cone on the counter electrode . As the droplets traverse the space between the needle tip and the cone and solvent evaporation occurs.
Once the droplet leaves the capillary and enters the nitrogen it continues to lose solvent until the charge density exceeds the surface tension, i.e. the Raleigh constant is exceeded, the droplet explodes resulting in smaller charged droplets. This process continues until the droplets are small enough for ion desorption. The creation of these ions facilitates the transfer of the sample molecules from the source into the MS as the ions are attracted and accelerated into the mass analyser.

Note: The first pioneering experiments on electrospray ionization (ESI) was conducted by Malcom Dole, Not me--- Malcolm. haha...

Guidelines for Choosing Ionization Method

Guidelines for Choosing Ionization Method for LC-MS

Note: This July, will focus on LC-MS and GC-MS@Pesticides.

So Do not go, I will be back --<(:) smile>- I learned this from FX TV channel, can not endure up to 10 times annoying TV Advertisements during a Movie... So I disconnected the cable services @ June-2008. Am I stupid? yes, maybe, but whatever....

Why 371,445 and 519 in my LCMS system?

So sweet question, today, remind me 2005 Fall, The First Question I asked at Tech.
All the optimal conditions were set up, where were they from?

One reasons could be they are from Contaminant Peaks from C18 Column Bleed. The contaminant ions, at M/Z 371, 445, and 519, when seen on the LC/MSD and LC/MSD Trap systems in positive ion mode, are believed to be C18 column bleed peaks of polysiloxane [O-Si(CH3)2-]n

Methylsulfonylmethane (MSM, or Dimethylsulfone)

Methylsulfonylmethane (MSM, or dimethylsulfone) is an organosulfur compound with the formula (CH3)2SO2. It occurs naturally in some primitive plants and is present in small amounts in many foods and beverages and it is marketed as a dietary supplement, although its benefits are disputed.

Purity and Limit of Dimethyl Sulfoxide ( DMSO) by Gas-Chromatography with FID.

Reference Standard: USP DMSO and MSM RS.
Column: 3m m x 0.5 mm capillary column coated with a 5 um phase G2.
Oven Temp: 120 degree C.
Inlet/ FID temp: 250 degree C.
Inlet Split ratio is 2:1.
He ( carrier gas) = 5 ml/min.
C-Air/H2 = 400:40 ml/min.

Result: MSM Purity is 101.03%, Dimethyl Sulfoxide ( DMSO) is 0.041%, Chromatography purity is 99.91%.

Good to go for Lunch.