Step 1.

Isolating
Penicillin Bacteria
Put piece of bread or citrus fruit in container in a dark place at 70 degrees F. It
should be in a closed (but not airtight) container. Add a few drops of water to the
container and leave all but one corner closed to keep in moisture.
It can take weeks for the mold to start growing. Hopefully you are making this in
advance of needing it.
The bread or fruit will start getting a gray mold. This mold will eventually turn a
bluish-green color. This is the mold that you want for making penicillin!
*You’ll want to use homemade bread.

Step 2. Making a
Host Culture for
the Penicillin to
Grow On
The greenish-blue mold you grew contains penicillin.
 “Natural penicillin” derived directly from mold is very sensitive to stomach acid. If
you drink it as a tea, it will be destroyed before it makes it to your bloodstream.
The only way to make this penicillin work is it to inject it and that is not good at
this stage because of the impurity and is not recommended!. To get the benefits of
penicillin from mold, it has to be further cultured.
You need to re-culture the penicillin. This step also ensures that you are only
getting penicillin bacteria and not salmonella.
 Re-Culturing
Penicillin
1. Thinly slice 200 grams of unpeeled potatoes.
2. Put the potatoes into a 1 liter mason jar and fill with distilled water. Screw on the
lid tightly. Put the entire jar into a pot of boiling water. Boil for 30 minutes.
3. Once it cools a bit, open the jar. Strain the contents through a cheesecloth . Catch
the liquid! This is what that you need.
4. Add 20g of glucose (dextrose) to the broth. If you don’t have glucose, plain sugar
will work.
5. .Add 20g of agar to the broth. If you don’t have agar, then plain gelatin (not Jello)
will work. Note that the agar isn’t going to completely dissolve.
6. Add distilled water until the total volume is 1 liter.
7. Pour the broth into wide, flat jars with a sealable lid. Basically, anything that could
work as a petri dish. If you actually have petri dishes, use them.
8. Cover the dishes immediately to prevent microbes from the air from getting into
them.

Step 3. Streaking
Your “Petri Dish”
Now it is time to move your penicillin mold spores to your “petri dishes” that have
the potato broth in them.
There is a very specific method that scientists use for growing cultures. It is called
“streaking.”
1. Get a thin piece of wire. Bend the tip into an oval shape.
2. Sterilize the tip of the wire in a flame. It should be red hot.
3. Dip the hot wire into the potato broth to cool it down (so the heat doesn’t kill your
penicillin spores).
4. Now touch the wire tip to the greenish-blue penicillin mold.
5. Make three lines on your petri dish. This is where the colonies of penicillin will
start growing.
Step 4: Let Your
Penicillin Grow
Keeping it covered, let the penicillin grow in your DIY petri dishes for about a
week.
Penicillin is a yellow color. However, other species of bacteria can also be yellow,
so it is hard to tell for sure that you got penicillin without a trained eye and a
microscope.

Step 5:
Fermenting Your
Penicillin
Here is where things get a bit complicated. You need to ferment your penicillin
spores so they reproduce in huge numbers. Otherwise, it will be like trying to put
out a forest fire with a bucket of water.
Most websites tell you to get a bunch of ingredients like potassium
monophosphate and sodium nitrate. That method does works – but I doubt you will
find those things laying around when SHTF.
Here is a simple method of fermenting penicillin.

Supplies
Erlenmeyer flask
(at least 500ml in size)
Graduated cylinder
Distilled water
Glucose (dextrose)
2g
 Yeast extract
1g
Citric acid
2g
Powdered nonfat milk
10g
Sea salt
1g
These ingredients are all available on Amazon.

Instructions
1. Sterilize the flask in the oven at 315 degrees F for one hour.
2. Put the glucose, yeast, citric acid, milk powder, and sea salt into your graduated
cylinder.
3. Fill the cylinder with water until it reaches the 100ml mark.
4. Pour the contents into your flask.
5. Put the lid on and shake until the contents are dissolved.
6. Add your penicillin cultures to the flask. Make sure you are using sterile methods,
such as using a sterilized metal loop to add the cultures!
7. Cover the flask with sterilized aluminum foil. This will keep unwanted microbes
out of the flask while still allowing for air flow.
8. Let the flask sit for at least 7 days but no longer than 14 days.Here is what
penicillin looks like growing in a glass flask.

Here is what penicillin looks like growing in a glass flask.

Step 6:
Extracting the
Penicillin
After about a week, you have got fermented penicillin in the flask. You've got to
get out all of the other ingredients before you can use it though.
 Supplies
Coffee filters
PH Tester
Hydrochloric acid

Instructions
1. There are going to be solid parts in the liquid. You need to separate these from the
liquid. The liquid is the part which contains the penicillin, so you want to hang on
to it!
2. Strain the penicillin liquid through a coffee filter or sterilized cheesecloth. Catch
the liquid in a sterile container.
3. Now you need to adjust the pH of the penicillin. Add a drop of hydrochloric acid
and then test the pH. It will probably be a pH of around 5 when you start. Keep
adding drops and testing until you get a pH of 2.2.
In theory, you could use this extracted penicillin now. However, the penicillin is
very unstable.
You’d have to use it right away. Considering that most infections need multiple
rounds of antibiotics to treat, your whole efforts would be for nothing.
Thus, you have to do the fairly complicated and scientific step of further extracting
the penicillin.

Step 7: Further
Extraction
Ethyl acetate is used to completely extract pure penicillin. The penicillin dissolves
into the ethyl acetate. Then the ethyl can be evaporated, leaving you with pure
penicillin.

Supplies
 Separator funnel
Ethyl Acetate
Potassium Acetate

Instructions
1. Chill ethyl acetate in a freezer.
2. Mix the cold ethyl acetate and penicillin liquid in a separator funnel.
3. Shake them together for 30 seconds, then allow them to separate again. The ethyl
acetate will sink to the bottom.
4. Open the separator funnel and allow just the ethyl acetate to slowly drip out into a
sterile collection container (a mason jar works for this).
5. Now add potassium acetate to the ethyl acetate. You’ll need about 1g of potassium
acetate for each 100ml of solution you have.
6. Leave the solution uncovered in a ventilated area. The ethyl will evaporate, leaving
behind penicillin.
You’ll note that we started with about 100ml of fermentation. From that amount,
you’ll probably only get about 100mg of penicillin.

Penicillium citrinum

Penicillium citrinum

Ecology: Penicillium citrinum is a commonly occurring filamentous fungus with worldwide

distribution. It has been isolated from a variety of sources including soils, decaying
vegetation, foodstuffs (beans, coffee, cereals & spices) as well as a variety of indoor
environments.

Pathology: While Penicillium species are generally regarded as laboratory contaminants, or at

best, opportunists, a number of species have been implicated as being involved in the disease
process. While Penicillium species may be isolated from clinical specimens, it is commonly
believed that a true infection can only be established by histological demonstration of tissue
invasion. With that in mind, Penicillium citrinum has been reported in mycotic keratitis (eye),
lung infections (pneumonia), a single case of a urinary tract infection (UTI) and one of
pericarditis. Their contribution to the disease process may be secondary an additional
underlying illness. As with all fungi, immunocompromised individuals may be at greater risk of
infection including those rarely considered as pathogenic.
 Macroscopic Morphology: Penicillium citrinum exhibits
moderately slow growth on Sabouraud-Dextrose agar (SAB) at 30ᵒC. Surface texture is
velutinous (soft, velvety surface) to floccose (woolly tufts of soft “hairs”). The colonial
growth appears radially sulcate (narrow, deep furrows or radial grooves –like spokes on a
wheel). The mature colony has a central greyish-turquoise to greyish-orange colour with a
white periphery (outer edge). Exudates (extrolites) are frequently produced which appear as
drops of liquid upon the surface of the colony. These may appear clear, to pale yellow, to a
reddish-brown in colour. Some strains may also produce a soluble pigment which can diffuse
into the surrounding medium. The reverse is a pale yellow to a light yellow-brown. Colours
and growth characteristics are, of course, media and strain dependent.

Penicillium citrinum-SAB, 14 days incubation at 30ᵒC (Nikon)

Note the drops of exudate (extrolites) which have formed on the surface.

Colour variation due to maturing of colony but also a difference in my lighting for
photography.
 Exudates (or Extrolites): Some fungi can produce exudates as a by-product of their growth,
many of which can be collected for commercial use. Mycotoxins are by-products (secondary
metabolites) which are potent poisons. Penicillium citrinum produces Citrinin, a nephrotoxic
mycotoxin which derives its name from the fungus. It may also produce other extrolites such
as tanzowaic acid A, quinolactacins, quinocitrinines, asteric acid and compactin.

Microscopic Morphology: Penicillium citrinum produces septate, hyaline (clear, not

pigmented) hyphae. Smooth-walled conidiophores stipes are rather long (100 – 300 µm) and is
biverticillate (see diagram at end of post). Metulae are 12 – 15 µm in length which are found
in whorls of 3 – 5 divergent structures. Phialides are ampuliform (flask-shaped) and about 7 –
12 µm in length. Conidia (2.2 – 3.0 µm dia.) are globose to sub-globose (round to off-round)
and are smooth or have a finely roughened surface. Conidia resist disruption and form rather
long chains. These characteristics: the metulae longer than the phialides and the conidia
being both spherical and produced in well-defined chains, are distinguishing features
of Penicillium citrinum.

Penicillium citrinum- distinguishing features of Penicillium 'species' can already be made out
at low magnification. (250X, LPCB, DMD-108)
 Penicillium citrinum- distinguishing features of Penicillium 'species' much more evident at
400X.

Typical "fingers" made up of the metulae and phialide structures from which chains of conidia
extend. (400X, LPCB, DMD-108)

Penicillium citrinum- a mass of overlapping fruiting structures with copious amounts of

conidia.

(1000X, LPCB, DMD-108)

Penicillium citrinum- a little less congested in this photo. Conidiophores (stipes) seen from
which extend the metulae and conidia producing phialides. Conidia are globose (round) to
sub-globose (somewhat off-round) in shape, (1000X, LPCB, DMD-108)

Penicillium citrinum- long metulae and the somewhat shorter phialides are clearly
distinguishable in this photograph. The conidia are generally smooth or can have a finely
roughened surface.

(1000+10X, LPCB, DMD-108)

 Penicillium citrinum- another view.

(1000+10X, LPCB, DMD-108)

Penicillium citrinum- exhibits biverticillate branching meaning that the conidiophore can
branch and the metulae & phialides extend from these branches. Triverticillate would have
the conidia branching and then the branches also branching to finally produce the metulae &
phialide fruiting structures.

(1000X, LPCB, DMD-108)

Penicillium citrinum- Phialides are ampuliform (flask-shaped) and about 7 – 12 µm in length.

Again, conidia (2.2 – 3.0 µm dia.) are globose to sub-globose (round to off-round) and are
smooth or have a finely roughened surface. (1000+10X, LPCB, DMD-108)
 Penicillium citrinum- Here we see the proportions of the metulae (M) and the 'flask-shaped'
phialides (P) with the metulae being substantially longer than the phialides, The biverticillate
structure is evident in this photo. (ie. each branch extending from the conidiophore (stipe),
branches only once and then bears a fruiting structure consisting of the metulae and phialides.

(1000+10X, LPCB, DMD-108)

Penicillium citrinum- another example.

(1000+10X, LPCB, DMD-108)

Penicillium citrinum- a few more photos to finish up.

(1000+10X, LPCB, DMD-108)

Penicillium citrinum- suitable for framing!

(1000X, LPCB, DMD-108)

 Penicillium citrinum

(400+10X, LPCB, DMD-108)

Penicillium citrinum

(400X, LPCB, DMD-108)

 Penicillium citrinum- another colony showing the exudate (extrolites) which accumulate on
the colony surface after extended incubation. These metabolites may be potent poisonous
mycotoxins or might have beneficial uses in industrial or pharmaceutical applications. (Nikon)

Physiology: The spores of Penicillium citrinum fail to germinate at 5ᵒC and may show
restricted growth at 37ᵒC.

* * *
Posted by Yuri at 18:28

Labels: Biverticillate, extrolites), exudates, metulae, Penicillium citrinum, phialides