Lecture 16 - Mycotoxins Flashcards
what are mycotoxins?
the toxins produced when molds (filamentous fungi) develops on food commodities)
what are hte main producers of mycotoxins?
fungal species belonging to the genera Asperigillus, Fusarium and Penicillin
what are the major classes of mycotoxins?
- aflatoxins
- ochratoxins
- trichothecens
- fumoisins and patulin
- zearalenone
what are examples of aflatoxins?
B1; B2; M1; M2; G1 and G2
what are examples of ochratoxins?
ochratoxin A
OTA
what are examples of trichothecens
DON (deoxynivalenol)
T2
HT2
what are examples of fumonisins
FBs: FB1, FB2, FB3
where are patulins found?
apples and apple products
what is the order of the food chain “farm to plate”
- preharvest farm
- harvest farm
- post harvest storage
- processing factory
- distribution transportation
- marketing wholesalers and retailers
- consumption
what are 2 categories of mold?
- field fungi
2. storage fungi
where are aflatoxins found?
sorghum soybeans corn wheat barley
where are trichothecens found?
barley oats sorghum soybeans corn wheat
where are zearalenone found?
wheat sorghum corn barley silage (forage grass, legume crops and corn)
what are the 3 methods of control and detoxification?
- physical methods: grinding and rinsing, heat treatment, irradiation, degradation, inorganic absorption
- chemical methods: alkalization, ozone degradation
- biological methods: microbial absorption, microbial degradation, biological degradation
examples of AFB1?
describe the properties of AFB1
- eg. aspergillus flavus and aspergillus parasiticus
solubility:
- soluble in polar organic solvents
- poorly soluble in water
- insoluble in petroleum ether, ethyl acetate and hexane
pH:
- stable in neutral solutions
- resistant to strong acids
- rapid decomposition in alkaline
UV:
- produces fluoresence (group B&G)
- destructive for low conc of AFB1
heat:
- stable in 200deg C
- decomposed until 268degC
- hard to destroy under normal temp
what are examples of Zearalenone ZEN
describe properties
eg. fusarium graminearum and fusarium culmorum
solubility:
- insoluble in water
- slightly soluble in hexane
- more soluble in benzene, acetonitrile, methylene, chloride, methanol, ethanol, acetone
- soluble in aqueous alkali
pH:
- stable in neutral and acidic
- ester bond will be open in alkaline environment
UV:
- exhibits blue-green
heat:
- melting point is 161degC
- hard to destroy under normal cooking
what are examples of deoxynivalenol?
describe properties
eg. fusarium and gibberella
solubility:
- soluble in water and polar solvents
- insoluble in hexane and diethyl ether
pH
- sensitive in alkaline environment
- stable in neutral and acid
UV:
- absorption peak under short wave UV
- decomposes under high tense UV
heat
- resistant to heat
- stable in 120degC for 1 hr
- decomposes at 150degC for 15 min
what are the types of pre-treatment/extraction in mycotoxin analysis
- liquid-liquid extraction (LLE)
- supercritical fluid extraction (SFE)
- solid phase extraction (SPE)
- solid phase micro-extraction (SIME)
what are 3 types of clean up detection in mycotoxin analysis?
- thin layer chromatography
- gas chromatography
- high performance liquid chromatography
how is a mycotoxin sampling plan defined?
by a mycotoxin test procedure and a defined accept/reject limit
how much can the number of containers sampled vary?
can vary from 1/4th (for small containers less than 20 metric tons) to the square root of the total number of containers (for large containers greater than 20 metric tons)
in sampling, how do you ensure that the analytical test portion has the same conc of toxins as the original sample?
the whole primary sample must be ground and mixed
describe the liquid-liquid extraction
- two immiscible phases shaken together
- compounds partition to their preferred phase (into aqueous or organic solvent)
describe the solid phase extraction/clean up and concentration
what are advantages?
- SPE is based on molecularly imprinted polymers that are syntehtic receptors for the mycotoxins
- used for sample extraction, clean up and concentration
- C18 (octadecylsilane), silica gel, anionic and cationic exchange materials, immunosorbets and molecular imprinted polymers
adv
- faster and most popular method
describe the solid phase concentration
what are advantages?
- uses immunoaffinity columns (IACs)
- used for purification of samples which are contaminated with different mycotoxins
- the mycotoxins (analytes) are bound selectively to the antibodies on the column
- rinsing removes most possible interferences. toxins can be eluted by antibody denaturation
adv:
- offers high selectivity
describe thin layer chromatography (TLC)
what gel is used?
what are advantages and disadv?
- used for detection step (for quantitative and semi-quantitative measurements of mycotoxins)
- detects by fluorodensitometry or visual procedures
- uses silica gel, F254 fluorescent silica gel, or silica gel impregnated with organic acid (for detection of aflatoxins, citrinin, fumonisin)
adv:
- costs less
- simple
- rapid screening
dis
- lack of automation has let to TLC being replaced by other techniques
describe gas chromatography for detection
- detection is achieved by linking the system to mass-spectrometry (MS), flame ionization or fourier transform infrared spectroscopy
why is GC not a technique suitable for commercial purposes?
not suitable for commercial purposes because of its limitations to volatile and thermostable compounds
when detecting mycotoxins with GC, why do mycotoxins need to be derivatised first?
what are examples of chemical reactions used for derivatisation?
since mycotoxins are not volatile
thus, derivitisation by chemical reactions (eg. silylation or polyfluoroacylation) are needed in order to be quantified
what is the most widely accepted official method for determination of mycotoxins?
HPLC
high performance liquid chormatography
describe detection by HPLC
HPLC = high performance liquid chromatography
what kind of detection is used in HPLC?
UV
fluorescence
amperometric
spectrofluorimetric
how can mycotoxins be detected directly by HPLC-fluorensence (HPLC-FD)
when mycotoxins have natural fluorescence (ochratoxin or citrinin)
what has increased detection sensitivity in comparison to HPLC-FD?
Mass spectroscopy with electrospray or pressure chemical ionization
what is thel imit of detection by HPLC-MS?
0.1 and 1 microg/Kg
what are advantages of HPLC?
disadv?
adv
- high resolution
- improved limit of detection
- possibility to be coupled to multiple detection automated systems
disadv:
- most chromatographic assays are expensive, time consuming and require expensive equipment and clean up
what is ELISA?
enzyme linked immuno sorbent assay
- based on the ability of a specific antibody to distinguish the 3D structure of a specific mycotoxin
- the direct competitive ELISA is commonly used in mycotoxin analysis
