I am trying to observe what would happen if I combined specific metals with distilled water, some salt and sugar with cocoa (low chlorine) present for a specific fungus (Penicillium I). It is known that brass, copper, zinc, and silver disrupt enzymes of fungi, cell wall and DNA and RNA but how different is PI (a mold) compared to specific bacteria and fungi (very different).
How will specific metals like Lead, Arsenic, and Copper effect the fungi that I am testing? I am testing a 1 million spore per ml sample (pretty high) but necessary for this project.
Silver and some other metals (Well especially silver) react to form insoluble inert silver chloride with chloride ions so low chloride ions for these test as well as distilled water is necessary.
WARNING: Some molds are pathogenic and dangerous to your health. Also, arsenic salts and lead salts if you try to make them are extremely toxic and even cancer-causing level 1. They can be fatal if absorbed through the skin, etc even at concentrations as low as 1%. Please do not ADD acids to lead, arsenic, etc if you are going to repeat this!! The metals are a LOT easier to handle than the salts.
Step 1: Prep Work for Plates and Metals and Mold on Plates Part 1
Prep work for agar.
Every 5 plates have 20 ml of cocoa agar. It consists of 100 ml approx distilled water, 5.0 grams sucrose, 5.0 grams Cocoa with sodium bicarbonate, and 5.0 grams Agar Agar. Heat in a hot plate for 15-20 minutes with occasional stirring until it begins to broth and foam up, then put on the flask on a cool surface. For a 100 ml sample of agar, mix make sure the flask is at least double the volume. Pour the hot agar 80 deg C into the plates and close them to avoid other contamination. Allow them to solidify.
Wearing gloves place each metal (Lead, arsenic) into the gloves. A bit of transfer of lead ions from metal will be transferred to the arsenic, etc but this is a minor and unavoidable problem.
Make a concentrated sample of water with mold samples and allow to stir without air for a few days and filter giving the active mold sample. Prepare this sample outside at 5 degrees C in open space (Not indoors) to avoid spore potential contamination!
Place 2 ml of concentrate mold sample onto agar cocoa plates. Close the plates.
Step 2: Prep of Silver Metal Over Copper With Other Types of Metals.
Here I will show how to get nanosilver (Near nanosilver particles) with copper and silver nitrate in distilled water.
In 20 ml of distilled water with 5 g silver nitrate add 6 g of copper metal.
Here the equation (Chemical to it).
2AgNO3 (aq) + Cu (s) >>>>> 2Ag(S) + Cu(NO3)2(aq).
5 grams silver nitrate/169.87g times 107.8682 g (Silver) = 3.17 g silver produced with 6 g copper. 2.85 g of copper left over in this reaction.
Warning: Pure Silver Nitrate solutions are toxic (well over 10 g per L) perm stain hands black with exposure and are corrosive to skin and eyes. Wear gloves and goggles to protect yourself if you work with silver nitrate.
Step 3: Results of Metals Part 1 (Brass, Lead, Arsenic, and Copper).
Here is the picture for the third entry here are some results for 6 days.
Brass---- Growth complete with Aspergillus Niger.
Lead--- Very little growth of A. niger.
Arsenic -----Very low growth of Aspergillus niger.
Copper pipe- the complete growth of black mold.
These indicate that arsenic and lead even though they are less active than copper and brass can strongly inhibit the growth of Aspergillus niger!! It may be due to Arsenic metal slowly releasing As+3 ions and Pb+2 ions for lead and their toxicity on human cells (disrupting enzymes including DNA, ATP, etc) even in small amounts may be why these metals have inhibitory effects on fungi.
However, growth did occur slower than brass and copper than arsenic and lead indicating it just slowed down the growth a bit (the mold was not completely killed by lead or arsenic metal). There some evidence that these types of fungi like some bacteria can pump out metal ions from the cell. In bacteria, the antibiotics can be neutralized or pumped out. This is a possible explanation for the slow mold growth on these plates.
Step 4: Silver (II) Oxide-silver Copper Powder, Zinc Powder, Copper Powder, Cobalt Powder and to Metal Fungi Test on Cocoa Agar.
First to make Ultra 99% pure silver(II)-oxide and silver powder with copper.
Add to 4 degrees C distilled water (Room temperature also will work--- but the particle size will be different effecting your results) and add 5 g 98% silver (I) nitrate to the water. Clean all beakers with distilled water since any tap water, etc, if you use it, would convert silver nitrate (due to chloride ions from chlorine) into silver chloride that would ruin the result totally.
Then add 10 g of 99.99% copper powder and stir well being careful to avoid splashes or too rapid stirring. (Lab coat, gloves, and goggles (Prefer Faceshield) is better since the silver nitrate stains badly skin and is fairly corrosive too.
Decant the cold water into another beaker and treat the leftover sol as trace silver nitrate (special disposal). Put in 50 ml of cold distilled water again and heat gently 50-60 degrees C on a Hotplate until very little water is Left. Let the rest air dry and place in a bottle or sealable bag for storage. Overheating may change silver metal into oxides by air exposure but Low heat can significantly reduce this problem.
Buy copper powder, silver powder and cobalt powder from e-bay.
Add 5 g of sugar to 100 ml tap water with 5 g cocoa 99%, and 5 g Agar-agar. If you are going to make 200 ml you obviously have to double the grams of sugar, etc to fit the same ratio. Boil the agar solution in a beaker double the volume of your sample or flask until some foaming and boiling happen ( A bit) then let it cool for 5-10 minutes. Be careful Agar turns into a gel at 50 degrees so do not wait too long since you cannot simply heat the agar back up into a liquid again (or very easily).
Let the plates that you poured 10 ml of each (20 ml if you have them) and let them solidify at room temp for 20 minutes approx.
Add 5 g by weighing and taring of a digital scale (approx 5 g) give or take 0.1 grams of the silver(II) oxide -silver metal with copper, copper powder, zinc powder, cobalt powder and To metal to each plate. Note that To metal is NOT a powder but 5 g is the amount I will use.
Step 5: Results for Part 2 (SIlver Oxide-Copper, Copper, Zinc and Cobalt, Etc
Here are the results for the following metal powders. Here the concentration
of each metal is 5 grams.
Silver (II) Oxide-Copper Powder----- Little growth for 2 weeks.
Copper Powder ------- Little growth for 2 weeks.
Zinc Powder ------ Little growth for 2 weeks.
Cobalt Powder -----Little growth for 2 weeks.
Step 6: Discussions and Conclusions.
Well, I am very surprised about the Metal powder. I did not think especially Silver (II) Oxide-Copper, Zinc or Cobalt was very effective at preventing the mold from spreading across the plate in 2 weeks!!! I think the size of the particles (Nano) allowed the metal to enter the mold thus destroying vital enzymes of the cell wall, DNA, etc.
I tried researching how metals like this kill Mold and I got nothing out of it. Bacteria it interferes with DNA synthesis, Cell wall reformation, RNA synthesis, etc. So I assume the metal is toxic in a similar manner but I could be wrong on that.
The metals (Lead, Arsenic, etc) surface area was too big to allow the metal to pass into the cell or enough of the metal inside to kill the Mold. However, powder including copper did.
However later On 1 month or so the Mold slowly grew right over the metals (Powder) indicating it only slows it down and just like the other metal it does not kill them.
2 Weeks of Metals like Arsenic, Lead, etc resulted in a complete growth of Black Mold (Aspergillus Niger) but the powder for 2 weeks results in some but it does not result in LAWN (Overgrowth of mold on the plate). The powder is far more effective at killing mold than regular metal is.
Eventually in a month lawn even over the metal powder grew over the agar.