Chromatography and Vet Drugs Flashcards by Nancy He

Why can’t we inject food samples directly into a detector? (2)

analyte is in very LOW concentrations

2. matrix interference

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What is a common sample preparation step for many toxin analyses? What is its purpose?

chromatography: to SEPARATE the analyte from the matrix

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Describe the basic function of chromatography:

separates components of a sample, distributing them between its MOBILE PHASE or STATIONARY PHASE

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chromatography is a ____ separation method.

physical

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What is GC? What analytes is it used for? (6)

gas chromatography:
used for organic compounds:
pesticides, POPs, PAHs, process induced contaminants, plasticizers, vet drugs + hormones

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What type of chromatography is used for inorganic substances?

Ionic chromatography: used for ions: sulfites, nitrates, cyanide

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What type of chromatography is mostly used for vet drugs? What else is it used for?

LC (liquid chromatography)

also for pesticides, POPs, process induced, plasticizers, vet drugs + hormones, mycotoxins

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What is used to seperate detergents?

ionic chromatography

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What is Tr?

retention time: time from sample injection to max elution peak (for compound of interest)

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What are the 4 mechanisms involved in chromatography? What is the basis for each?

adsorption - affinity
sieving - size
ion exchange - charge
partitioning - hydrophobicity/solubility

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The (greater/less) the affinity, the longer the retention time.

greater

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The (larger/smaller) the particle size, the longer the retention time in size exclusion chromatography.

smaller

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Describe the basic parts of of GC system.

long COLUMN Inside column oven (coiled) is stationary phase
CARRIER GAS is supplied, through FLOW CONTROLLER (mobile phase)
sample injected -> carried by gas through column
travel to DETECTOR -> generate SIGNAL

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What are some common GC detectors? (6)

Flame ionization (FID)
Nitrogen-Phosphorus (NPD)
electron capture (ECD)
MS
tandem MS (MS/MS)
Hi-res MS (HRMS)

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What GC detector can be used for all organics?

FID

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what effect does increasing temperature have on GC?

increase volatility of compounds -> faster process

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What can be done do separate compounds based on volatility in GC?

temperature gradient: start high, then lower (volatiles eluted out first)

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FID is ___ but not ____. why?

sensitive; selective (lack specificity)

based on burning; many compounds in food are C containing.

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NPD is good for:

Why? What might be a problem?

carbamates (pesticides)
contain N (reacts w/ N)
problem: compounds in food also can have N or P (interference)

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Which of the GC detectors are accepted for REGULATORY PURPOSES?

MS, MS/MS, HRMS

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What is the basis of the ECD in GC and what is it good for? Why might this be problematic?

react w/ electronegative atoms (Cl, Br): good for OCPs, PCBs

food can also have electroneg atoms (interference)

What detector is used for dioxins?

HRMS

POPs should be analyzed with:

MS or MS/MS

PAHs should be analyzed with:

how does LC system differ from GC? (4)

1. mobile phase is LIQUID SOLVENT 2. delivered through PUMP 3. stationary phase is HPLC column - SOLID PHASE 4. use different DETECTORS

why is a pump necessary for LC?

require PRESSURE to push liquid through solid phase (HPLC column)

What are the detector types for LC? (6)

``` UV-Vis Spectrophotometer diode-array (DAD) fluorescence (FLUO) conductivity (COND) MS/MS HRMS ```

Pesticides are used with what types of LC Detectors? (4)

UV-vis, DAD, MS/MS, HRMS

What is the basis of UV-vis detectors and DAD?

UV vis: based on ABSORPTION (measure @ 1 wavelength) | DAD: also absorption, but many wavelengths

What LC detector is used for aflatoxins, and what is its basis of detection?

FLUO: detect fluorescence (absorbing and emitting light @ diff wavelength)

COND is used for:

ions (cyanide)

the data generated by GC or LC is called a:

chromatogram

How are compounds identified through GC or LC? (2)

1. retention time (or relative retention time, comparing to standard) 2. criteria specific to detector (fluorescence, max absorbnce, exact mass, etc) * need to match BOTH CRITERIA

True/False: we can identify a compound based on its retention time only

False; compounds may have same retention time, need to also observe detector criteria (exact mass, etc)

What should be done to increase accuracy of the detector?

proper sample preparation; remove interferences, concentrate analyte

The size of the peak on a chromatogram depends on:

1. concentration | 2. sensitivity to analyte (some produce bigger signals)

How can a chromatogram peak be used to quantify the analyte?

integrate area under curve | compare to standard

the 2 types of vet drugs:

1. growth promoters - increase growth | 2. antibiotics - treat/prevent disease

What is the major antibiotic type? What are some others used?

TETRACYCLINES | macrolides, penicillin, lincosamides, aminoglycosides, sulfas

What are the "not medically important" vet drugs?

growth promoters: ionophores & NIR

most drugs are administered by: | what other methods are possible?

feed | water, injection, intramammary, oral/topical

What are the concerns related to vet drug use?

RESIDUES in food! 1. acute effects (allergy, toxicity) 2. long term effects (damage reproductive health, carcinogen, mutagen) 3. impact on gut microflora: kill 'good' gut microbes; INCREASING BACTERIAL RESISTANCE!

True/False: there is a higher incidence on non-compliant vet drug residue levels in imported animal products

False; not for all animal products (sometimes domestic is higher)

What carcinogenic compound was used in aquaculture? why? is it still used today?

MALACHITE GREEN antifungal/anti-protozoa properties; cheap and effective PROHIBITED; but still found in some imported seafood

analysis of vet drugs follows a similar procedure as: | What are some key differences?

POPs but: more POLAR (use polar solvents)

Describe the basic procedure for analysis of vet drugs:

1. preparation: homogenize and dry (oven or freeze-dry) 2. extraction: SPE or solvent (LPE) - ethanol, acetone 3. clean-up: column separation (or SPE) 4. concentration: rotary or N2 evap 5. analysis (LC-UV, LC-FLUO, LC-MS/MS, GC-MS, GC-MS/MS)

what procedure can accomplish both extraction and clean-up?

SPE (solid phase extraction)

what are the analysis machine methods used for vet drugs? (5)

``` LC-UV LC-FLUO LC-MS/MS GC-MS GC-MS/MS ```

the standard/official methods for vet drug analysis are:

HPLC-MS/MS | HPLC-HRMS

vet drug residues are usually found in what foods, and in what concentration range?

animal origin foods (meat, fish, eggs, milk) | ppb-ppm

What is the preferred separation method for vet drugs analysis?