Finding a calming molecule in coastal seawater bacteria
2 were GABA producers
optimised one at a time
reached at 48 hours
unoptimised starting medium
A laboratory study of marine bacteria from the Visakhapatnam coast that make GABA, the brain’s main inhibitory neurotransmitter. It covers how the bacteria were found and identified, how their output was nearly doubled, and how GABA was separated from the broth.
Methods and tools used
Bench techniques to isolate, identify, optimise and purify GABA producers, and the tools used to analyse sequences and write up the results.
- Aseptic isolation and culture
- Thin-layer chromatography
- Biochemical identification
- Antibiotic susceptibility testing
- Fermentation optimisation
- Spectrophotometric assays
- Ion-exchange chromatography
- 16S rRNA sequence analysis
- BLAST and BioEdit
- Scientific writing
Why this project matters
GABA is used in supplements, functional foods and medicines for stress, sleep and blood pressure. Fermentation by bacteria is a clean way to make it, and the sea is a largely unexplored place to look for new producers.
The problem
Most GABA research uses lactic acid bacteria from fermented food. Marine bacteria live under salt and pH stress, and in many bacteria GABA production is part of how they cope, so they may be strong producers.
Very few studies have isolated, optimised and purified GABA from bacteria on the Indian coast, so there is little practical data to build on.
The approach
Collect seawater, isolate bacteria, screen for GABA, identify the producers by DNA sequencing, then tune the growth medium step by step and test a simple purification.
Safety checks on blood agar and an antibiotic panel were included to judge how the strains could be used responsibly.
Three questions the study set out to answer
Discovery
Do bacteria from coastal seawater make GABA, and which species are they?
Optimisation
Which sugar, nitrogen source, pH, temperature, mineral, vitamin and substrate level give the highest yield?
Purification
Can GABA be separated from leftover glutamate in the fermentation broth?
From the shore to the data
Each stage answers one of the research questions and feeds the next.
Isolation and
screening
Seawater and fermented batter were diluted and plated, and pure colonies were grown in glutamate-rich medium. Culture supernatants were run on thin-layer chromatography plates and compared with a GABA standard.
Strain
identification
Colony shape, Gram staining and seven biochemical tests described each isolate. The 16S rRNA gene was sequenced and matched against the NCBI database to name the species.
Safety
profiling
Blood agar showed whether the strains destroy red blood cells, and a seven-antibiotic disc panel showed how resistant they are.
Fermentation
optimisation
Carbon source, nitrogen source, pH, temperature, metal ions, vitamin B6 and glutamate level were each varied in turn, keeping the best result before moving on. GABA was measured at 24, 48 and 72 hours in triplicate.
Partial
purification
Broth was cleared, decolourised, desalted and passed through an anion-exchange column. Flowthrough, washes and acid elutions were checked by chromatography.
I’d be glad to talk through my findings
My work is yet to be published, so I’m not sharing the detailed results here just yet. If you’re interested, please get in touch and I’ll happily walk you through what I found.
- Emailkvh720@gmail.com
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