Phytotoxin
phtotoxin produce by bacteria and fungi
Bacterial toxin are two types endotoxin and exotoxins
Fungi produce toxin Mycotoxins
Mycotoxins - Aflatoxins B1, B2, G1, G2,
Products contaminated by aflatoxins such as cereal, tree nuts, dry fruits, spices, dairy products, eggs, and medicinal plants.
There are various methods use for the detections of aflatoxins like HPLC, HPTLC, ELISA,TLC, and LC-MS.
Aflatoxins cause chronic and acute toxicity.
Chronic- slow growth, immunity problems, cirrhosis, and liver cancer.
Acute- Hemorrhage, edema and acute liver toxicity.
2. PHYTOTOXIN
A poisonous substance derived from a plant.
A substance that is phytotoxic, especially one
produced by a parasite.
phytotoxin are members of various classes
of secondary metabolites,
including alkaloids, terpenes and especially phenolics,
though not all such compounds are toxic.
Phytotoxins may also be toxic to humans
3. PRODUCTION OF PHYTOTOXIN
Produced by bacteria
Produced by fungi
The toxins may be glycosides, amino acid
derivatives, peptides, terpenoids, sterols,
pyridines, and quinones, etc.
BACTERIAL TOXINS
Two types of bacterial toxins
Endotoxin
Exotoxin
4. CHARACTERISTICS OF BACTERIAL ENDOTOXINS
AND EXOTOXINS
PROPERTY ENDOTOXIN EXOTOXIN
Chemical nature Lipopolysaccharide (mw
= 10kDa)
Protein (mw = 50-
1000kDa)
Relationship to cell Part
of outer
Part of outer membrane Extracellular, diffusible
Denatured by boiling No Usually
Antigenic Yes Yes
Form toxoid No Yes
Potency Relatively low (>100ug) Relatively high (1 ug)
Specificity Low degree Highdegree
Enzymatic activity No Usually
5. TOXINS PRODUCED BY FUNGI
Mycotoxins Main Producing Fungi
Aflatoxins B1, B2, G1, G2 Aspergillus flavus, A.
parasiticus, A. nomius
Ochratoxin A A.alutaceus, A.carbonarius
Patulin P. expansum, A. clavatus,
Fumonisins Fusarium proliferatum
TYPES OF MYCOTOXINS
Mycotoxins = Toxic metabolites of fungi.
Mycotoxins contaminate the wide variety of food
as a result of fungal infection in crops, during
growth or in storage.
6. AFLATOXINS
The name Aflatoxin comes from
A (Aspergillus)
FLA (flavus) toxin.
Aflatoxins are a group of structurally related toxic secondary
metabolites produced by three species: Aspergillus flavus,
Aspergillus parasiticus and the rare species A. nomius.
It known to be highly toxic and potential carcinogens.
Aflatoxins were first identified in 1961 in animal feed
contaminated by Aspergillus parasiticus
7. CONTD…
Aflatoxins are the only mycotoxins currently regulated by the
U.S. Food and Drug Administration.
According to International Agency for Research on
Cancer(IARC) In 1988, the aflatoxin B1 list of human
carcinogens.
Studies have shown that concurrent infection with the Hepatitis
B virus (HBV) during aflatoxin exposure increases the risk of
hepatocellular carcinoma
Aflatoxins B1 and B1 = blue fluorescence
Aflatoxins G1, G1 = yellow-green fluorescence
8. MAJOR TYPES AND THEIR METABOLITES
Aflatoxin B1 is considered the most toxic and is produced by
both Aspergillus flavus and Aspergillus parasiticus.
Aflatoxin M1 is present in the fermentation broth of Aspergillus
parasiticus, but it and aflatoxin M2 are also produced when an
infected liver metabolizes aflatoxin B1 and B2.
Aflatoxin B1 and B2, G1 and G2, produced by Aspergillus
flavus and A. parasiticus
Aflatoxin M1, metabolite of aflatoxin B1 in humans and
animals
Aflatoxin M2, metabolite of aflatoxin B2 in milk of cattle fed
on contaminated foods
11. CHEMICAL STRUCTURE
Aflatoxins are normally refers to the group of
difuranocoumarins and classified in two broad groups according
to their chemical structure.
A. Difurocoumarocyclopentenone series (AFB1, AFB2 )
B. Difurocoumarolactone series (AFG1, AFG2,).
12. DETECTION IN HUMANS
There are two principal techniques that have been used to detect
levels of aflatoxin in humans.
The first method is measuring the AFB1-guanine adduct in the urine
of subjects. The presence of this breakdown product indicates
exposure to aflatoxin B1 during the past 24 hours.
Another technique that has been used is a measurement of the
AFB1-albumin adduct level in the blood serum. This measure of
exposure over several weeks or months.
METHOD OF ANALYSIS
ELISA technique (Enzyme linked immunosorbent assay)
TLC (Thin Layer Chromatography)
HPLC - FLD (High Performance Liquid Chromatography)
HPTLC (High Performance Thin Layer Chromatography)
LC-MS
13. THIN-LAYER CHROMATOGRAPHY (TLC)
Thin-layer chromatography is one of the most widely
used separation techniques in aflatoxins analysis. It
consists of a stationary phase made of either silica or
alumina or cellulose immobilized on an inert material
such as glass or plastic, called the matrix.
The mobile phase is comprised of methanol :
acetonitrile : water mixture
which carries the sample along as it moves through the
solid stationary phase. In TLC, the distribution of
aflatoxins between the mobile and stationary phases is
based primarily on differences in solubility of the
analytes in the two phases.
14. CONTD…
Different analytes, depending on their molecular
structures and interaction with the stationary and
mobile phases.
Thin-layer chromatography has been widely used in
the determination of aflatoxins in different foods] and
as low as 1–20 ppb of aflatoxins has been reported .
The advantage of using the TLC method is that it can
detect several types of mycotoxins in single test.
Whereas TLC has excellent sensitivities
15. CONTD…
In addition, TLC accumulated errors during sample
application, plate development, and plate
interpretation. Attempts to improve TLC have led to
the development of automated form of TLC, called
the High-performance thin-layer chromatography
(HPTLC).
The HPTLC has since overcome the problems
associated with the conventional TLC techniques
through automation of sample application,
development, and plate interpretation.
The currently HPTLC is one of the most efficient and
precise methods in aflatoxins analysis.
16. HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY (HPLC)
High-performance liquid chromatography (HPLC) is the most
popular chromatographic technique for separation and
determination of organic compounds.
The HPLC technique makes use of a stationary phase
confined to either a glass or a plastic tube and a mobile phase
comprising aqueous/organic solvents, which flow through the
solid adsorbent.
The sample to be analyzed is usually injected into the
stationary phase and the analytes are carried along through the
stationary phase by the mobile phase using high pressure
delivered by a pump. The analytes are distributed differently
within the stationary phase through chemical as well as
physical interactions with the stationary and mobile phases.
17. CONTD…
The time is recorded by a detector as its retentionThe
retention time depends on the nature of the analyte and
composition of both stationary and mobile phase.
Programmable detectors such as either the fluorescent
detector (FLD) or the ultraviolet (UV) detector or the
diode array detector (DAD) may be used in the
detection and identification of aflatoxins.
ENZYME-LINKED IMMUNOSORBENT ASSAY
(ELISA).
The preparation of enzyme-antigen conjugates and
enzyme-antibody conjugates by Avrameas in 1969 for
the enzyme-linked immunosorbent assay (ELISA)
development.
18. CONTD…
This is methods of choice for medical diagnostic
laboratories, research institutions, andregulatory
bodies for quality assessment and proficiencytesting,
among others.
The ELISA technique is currently used in the
detection of aflatoxins in agricultural products and a
number of commercially available ELISA kits based
on a competitive immunoassay format are widely
used.
Most of the kits use horseradish peroxidase (HRP)
and alkaline phosphatase (AP) enzymes as labels in
analysis of aflatoxins.
19. CONTD…
TYPES OF ELISA
INDIRECT ELISA DIRECT ELISA
SANDWICH ELISA COMPETETIVE ELISA
MATERIALS NEEDED FOR ELISA KIT
ELISA Plate
Positive control
Negative control
Dilution Buffer
Conjugate
TMB Substrate
Stop Solution
21. TOXICITYOF AFLATOXIN
Acute aflatoxicosis
High dosage over short time
Hemorrhage
Acute liver damage
Edema
Altered digestion, absorption, and
metabolism
Death
Chronic aflatoxicosis
impaired food conversion
slower growth
immunity problems
cirrhosis
liver cancer
22. PRE-HARVEST RISK FACTORS
High temperatures
Heavy rains
Crop insect damage
Poor fertility
Weed competition
High crop densities
Post-harvest risk factors
High temperatures
Humidity
23. TOXIN PRODUCE BY PLANT
MASHROOM
There are many symptoms observed in humain
beings ingestion for mashroom.
24. DATURA
FAMILY : Solanaceae
POISONOUS PART : All parts
POISONOUS CONTENT : Tropane alkaloids
SEVERE POISONING
Disorientation
Hallucinations
Respiratory depression
Coma
TREATMENT
Induce emesis or gastric lavage
Catheterization to empty bladder if necessary
Diazepam for hallucinations
25. CANNABIS SATIVA
FAMILY: cannabaceae
POISONOUS PART: All parts
POISONOUS CONTENT:
canabiol, canabinolic acid, canabinoids
TOXICITY
Tachycardia
Increased systolic pressure
Cough
Hallucinations
Confusion
TREATMENT: Intoxication within 4-6 hrs. In case of overdose
medical supervision is needed.
26. NUX-VOMICA
Family : Loganaceae
Poisonous part : seed (although all parts are toxic)
Poisonous content : Strychine, brucine
Toxicity:
Symptoms appear within 15-30 mins of ingestion
Suffocation
Extreme contractions of muscles in the body
Renal failure
Death is caused by muscular paralysis
Treatment:
Activated charcoal
Support respiratory and cardiovascular functions
When convulsions and hyperactivity are completely
controlled, gastric lavage can be performed safely
