Jashore University of Science and Technology

Course Title: Food Safety and Toxicology

Course code: NFT-5209

Presented by Presented to
Md. Riazuddin Professor Dr. Shireen Nigar
Roll: MS-210931 Chairman
Dept. of Nutrition and Food Dept. of Nutrition and Food
Technology, JUST. Technology, JUST.
Inauguration

Good morning, Greetings to all of the attendees..

Welcome to my presentation.

Title: Toxicological and biochemical responses of the earthworm

Eisenia fetida to cyanobacteria toxins.
Background Eutrophication

 What is the eutrophication of water?

The process in which a water body becomes overly
enriched with nutrients leads to the plentiful growth
of simple plant life.
 Indicators of Eutrophication in water?
The excessive growth (or bloom) of algae and
plankton in a water body are indicators of this
process.
 Causes of Eutrophication
The availability of nitrogen and phosphorous limits
the growth of plant life in an ecosystem.
These nutrient-containing water bodies favor the
growth of Algae, plankton, and other simple plant
life over the growth of more complex plant life.
Background Eutrophication

 Steps of eutrophication Nutrient

enrichment that
occurs due to
runoff from
agricultural fields

Aquatic species die

as a result of the Rapid growth of
loss of oxygen and algae and other
the production of plankton resulting
dangerous poisons. in an algal bloom

Dissolved
oxygen depletion Algae die and are
and toxin decomposed by
generation. bacteria
Background Cyanobacteria and their cytotoxins

Cyanotoxins can be produced by a wide variety of

planktonic cyanobacteria.
Some of the most commonly occurring genera are
Microcystis, Dolichospermum, and Planktothrix.

The following toxins are produced by cyanobacteria.

 Neurotoxins (Anatoxins, Domoic acid, saxitoxin)
 Hepatotoxins (Microcystin)
 Cytotoxins
 Dermatotoxins
 Irritants toxins.
Background Earthworm and Cytotoxins

 Role of earthworms in soil fertility

 Improved nutrient availability and cycling
 Improved drainage
 Water purification
 Remediation
 Restoration
 Improved productivity

 Impact of cytotoxins on earthworms

 Reduced lifespan
 Extended generation time
 Decreased brood size
 Delayed Development
 Suppressed locomotion
Materials and methods Extraction and analysis of microcystin variants

Freeze Ultrasonic
drying Homogenization bath (5min) Centrifuge at
Oasis HBL 25 mL, 5% 10,000 r/min for
extraction Acetic acid ) 15 min

Re-extraction
Cyanobacterial
Fig: Flow Diagram
bloom extract Cartridge with toxin
rinsed (50 mL, 5%
methanol)

MC-LR
MC-YR Addition of 10 Evaporation Elution
UPLC-MS/MS mL of
MC-RR (100 mL, 100%
deionized water methanol)
Materials and methods Avoidance test

Collection of Homogenization
soil sample and separation

Mixing of MC
deionized water

Materials and methods

Placing earthworms
between control
and sample soil

Incubation at 20
degrees for 48 h

NR=(C-T)/N×100%
Materials and methods Analysis of MCs in earthworms

Homogenization Ultrasonicatio
with 10mL of n Centrifugation
(BuOH:MrOH:H2O)

Grinding Supernatant
mixing (20mL,
5% alcohol)
Lyophilized/
Freeze-drying Evaporation

Elution
UHPLC/MS (15 mL, 100%
Stored at -40C methanol
Materials and methods Analysis of MCs in Artificial soils

Lyophilized soil Ultrasonication

extraction Homogenization
30mL 0.1M EDTA
Lower pH 3
with TFA

Centrifugation

Lyophilization
of soil Elution

Stored at -40 C
UHPLC/MS Evaporation
Materials and methods Biochemical assays

Earthworm
grinding in Centrifugation Homogenization Supernatant
0.85% NaCl s

Catalase (CAT) assay kit

Catalase activity
(Ammonium molybdate method)

Glutathione peroxidase (GPx) assay

GPx Activity
kit (Colorimetric method)

Total superoxide dismutase (T-

Total SOD activity SOD) assay kit (Hydroxylamine
method)

Total protein content Total protein assay kit

Result Effect
Biochemical
of MCs assays
Result Effect of MCs on Lysosomal membrane stability
Result Results of ANOVA tests

Figure: ANOVA tests of the biochemical response of earthworms exposed to MCs. The
dose or the duration of exposure had a significant effect (*p<0.05, **<0.01). df, degree
of freedom.
The end

Thank you for your patience and

attention.