Project - Electrical Stimulation on Eradicating Bacteria/Biofilms

Hello all,

I have a project in mind, but I am fairly new to project design.

The goal is to produce a device similar to a modified TENs device. The idea would be to have controllable parameters such as voltage, current, and wave forms.

The research I have on eradicating bacteria/biofilms all range from either saying pulsed high voltage works well, to direct current varying from 20 uA to 2,000 uA works well, to a combination of both with varying application time.

I am not interested in which works the best as I will perform my own experiments. I just need help in understanding what could be develop for an electrical stimulation output device with controllable/adjustable parameters using Arduino.

Thank you. I have also attached some interesting articles on the matter for reading.

Biofilm ES Current.pdf (640 KB)

Is this for lab research using machines, or for human subjects? The Arduino processor is specifically excluded from medical use.

aarg:
Is this for lab research using machines, or for human subjects? The Arduino processor is specifically excluded from medical use.

Lab research using ES on various bacteria and biofilm.

You will need to design and build the device that generates the voltage and current required, and it will need some way of being controlled by 5V logic signals.

By comparison, whatever the Arduino does will be completely trivial.

Are you a penniless researcher, or a paid professional working for a commercial venture?

jremington:
You will need to design and build the device that generates the voltage and current required, and it will need some way of being controlled by 5V logic signals.

By comparison, whatever the Arduino does will be completely trivial.

Right, and this is what I imagined using Arduino would be implemented for in this project. I didn’t think Arduino could be capable of acting as the main power supply for the system, but rather Arduino would function more as a programmable controller, for changing between various settings such as voltage, current, and frequencies levels.

If I have a circuit design to output a max of 100 V, or 1 A of current, is there a way to use Arduino to select voltage/current control to adjust the value of voltage or current between 0 and it’s max? I’ve managed to artificially out voltage levels between 0 - 5 V using a keypad and measuring with a multi-meter, but I realized once I connected the Arduino output pin to an oscilloscope I am really changing the duration on/off state of the pin.

Here is the code:

// include the library code:
#include <LiquidCrystal.h>
#include <Keypad.h>

// initialize the library with the numbers of the interface pins
LiquidCrystal lcd(A0, A1, A2, A3, A4, A5);
const byte ROWS = 4; //four rows
const byte COLS = 4; //four columns
char keys[ROWS][COLS] = {
  {'1','2','3','A'},
  {'4','5','6','B'},
  {'7','8','9','C'},
  {'*','0','#','.'}
};

byte rowPins[ROWS] = {5, 4, 3, 2}; //connect to the row pinouts of the keypad
byte colPins[COLS] = {9, 8, 7, 6}; //connect to the column pinouts of the keypad



Keypad keypad = Keypad( makeKeymap(keys), rowPins, colPins, ROWS, COLS );

// Global variables
byte index = 0;
char numbers[20]; // Plenty to store a representation of a float


long outPut = 10;      // LED connected to digital pin 9


void setup()
{
  lcd.begin(16, 2);
  // Print a message to the LCD.
  lcd.print("Setting up...");
  delay(1000);
  lcd.clear();
  lcd.print("Initializing");
  lcd.setCursor(0,2);
  lcd.print("Voltage..."); 
  delay(5000);
  lcd.clear();
  lcd.print("Voltage Set.");
  
  delay(1000);
  lcd.clear();
  lcd.print("Ready to use.");
  delay(1000);
  lcd.clear();
 
  
 Serial.begin(9600);
 pinMode(outPut, OUTPUT);   // sets the pin as output
 
 delay(500);
 lcd.clear();
}

void loop()
{
 
   char key = keypad.getKey();
  
   if(key != NO_KEY)
   {
       Serial.println("Key Pressed: ");
       Serial.print(key);
       Serial.println();
       lcd.print(key);
       
    
       
      if(key == 'C')
      {
         index = 0;
         numbers[index] = '\0';
         lcd.clear();
      }
      else if(key == '.')
      {
         numbers[index++] = '.';
         numbers[index] = '\0';       
      }
      else if(key >= '0' && key <= '9')
      {
         numbers[index++] = key;
         numbers[index] = '\0';
      }
      else if(key == '#')
      {
         float len = atof(numbers);
         long voltage = (1023/5) * len;
         long voltage2 = voltage / 4;
         
         // Do whatever you need to with len
         if(len > 5)
         {
           lcd.clear();
           lcd.print("Error!");
           lcd.setCursor(0,1);
           lcd.print("Voltage reset.");
           analogWrite(outPut, 0);
         }
         else
         {
         Serial.print("Numbers: ");
         Serial.println(numbers);
         Serial.print("Len: ");
         Serial.println(len);
         lcd.clear();
         lcd.setCursor(0,1);
         lcd.print("Voltage: ");
         lcd.print(len);
         lcd.print("V");
         
         voltage = (1023/5) * len;
         voltage2 = voltage / 4;
         analogWrite(outPut, voltage2);
         
         index = 0;
         numbers[index] = '\0';
         lcd.setCursor(0,0);
         lcd.print("Entered: ");
         lcd.print(numbers);
         }


      }
   }
}

aarg:
Are you a penniless researcher, or a paid professional working for a commercial venture?

I am a penniless researcher.

If I have a circuit design to output a max of 100 V, or 1 A of current, is there a way to use Arduino to select voltage/current control to adjust the value of voltage or current between 0 and it's max?

Of course. You can buy such programmable power supplies, or hire/collaborate with someone who has quite a bit of experience to design and build it.

Neither approach will be cheap or simple.

TGH904:
I am a penniless researcher.

You might find power supplies on the used market that can be controlled via a serial interface. I think HP made some.

jremington:
Of course. You can buy such programmable power supplies, or hire/collaborate with someone who has quite a bit of experience to design and build it.

Neither approach will be cheap or simple.

aarg:
You might find power supplies on the used market that can be controlled via a serial interface. I think HP made some.

I am compiling together an excel spreadsheet of all the various in vitro experiments that have already been done using electrical stimulation and I am coming across a wide range of used parameters, which is fine like I said, but I have a basic electrical engineering question I am wondering about with using one of these programmable power supplies.

If one researcher states that direct currents of anywhere between 500 uA to 5 mA successfully kills bacteria, and another states that high pulsed voltages around 250 - 300 Volts at 40 - 128 pulses per second successfully kill bacteria. Can I combine these values and stimulate bacteria with a pulsed 250 - 300 Volts AND have a current flowing in the range of 500 uA - 5mA?

In my head I almost imagine these pulses acting like a switch, and when they are switched 'on' current is also being the circuit element (bacteria).

Thanks again for all your help.

@aarg and @jremington

TGH904:
I am compiling together an excel spreadsheet of all the various in vitro experiments that have already been done using electrical stimulation and I am coming across a wide range of used parameters, which is fine like I said, but I have a basic electrical engineering question I am wondering about with using one of these programmable power supplies.

If one researcher states that direct currents of anywhere between 500 uA to 5 mA successfully kills bacteria, and another states that high pulsed voltages around 250 - 300 Volts at 40 - 128 pulses per second successfully kill bacteria. Can I combine these values and stimulate bacteria with a pulsed 250 - 300 Volts AND have a current flowing in the range of 500 uA - 5mA?

In my head I almost imagine these pulses acting like a switch, and when they are switched 'on' current is also being the circuit element (bacteria).

Thanks again for all your help.

@aarg and @jremington

your imagination is correct. The bacteria HAVE to be part of the circuit. They must have some resistance value or there will be no voltage pulse applied to them. No resistance means no measurable voltage, but huge current.

Your challenge will also be to attach electrodes to the bacteria.

Or are the past experiments only using the electric field, like between plates of a capacitor, to cause an effect on the bacteria?

Paul

Paul_KD7HB:
your imagination is correct. The bacteria HAVE to be part of the circuit. They must have some resistance value or there will be no voltage pulse applied to them. No resistance means no measurable voltage, but huge current.

Your challenge will also be to attach electrodes to the bacteria.

Or are the past experiments only using the electric field, like between plates of a capacitor, to cause an effect on the bacteria?

Paul

That is a good point. The papers I have read have either said "I am applying direct current at variable x uA/mA intensity directly to the surface of the bacteria", or "I am pulsating high voltages directly to the surface of the bacteria". Yet with their experimental design(s) I never understood how exactly they are:

  1. Measuring said values across their load (the bacteria)
  2. Actually applying these values to their load (bacteria) based on their setups

I will attach some articles for anyone to view if they want a better understanding of some of these other experimental setups, and if anyone understands them differently than I do in the electrical engineering world of things, please let me know. I may have completely misinterpreted their experimental design(s).

I have already tried an experiment a few times before using a TENs device. I just wanted to see if I could 'kill' or reduce the amount of bacteria with a TENs unit and store bought electrodes.

What I found was the electrodes killed more bacteria then the pulsating TENs voltage did. To my understanding it did this simply because I had "suffocated" the bacteria under the surface of the pad material, and the material was naturally antimicrobial. I was using a pulsating setting around mid range intensity, which I measured on an oscilloscope to be about 50 V, 5.0 Hz, 100 uS Pulse Width. I reason this because bacteria still grew/existed all around the pads (outlined them to be exact), but underneath each pad (anode/cathode) was a clear indication I had killed anything under the pad as nothing existed there. I hope this made sense.

I have revised my experiment to hold a coupon (a tiny disk covered in bacteria) suspended by tiny plastic pillars between two electrodes at fixed distances apart from the coupon, with a medium flowing between them to complete the connection (basically a salty solution). Similar to the one in the attached article titled "Direct Electric Current Treatment under Physiologic Saline Conditions Kills Staphylococcus epidermidis Biofilms via Electrolytic Generation of Hypochlorous Acid". Note* I have not tried this experiment yet, therefore I do not know the outcome.

Link to previous experiment using a similar experiment
Link to Direct Current Experiments
Link to four different electrical stimulation experiments

I do not expect anyone to read these experiments fully, but if you scroll to where they mention any experiment setup, or any images of their experiment, maybe you can then say something about what I need to do to replicate a similar experiment using Arduino/Power Supplies etc.

I made a block diagram to help me outline what I need to do to build a power supply similar to theirs and things may need to be switched around, but let me know what you all think. Thanks again.

Power Supply-> Voltage Regulator -> Voltage Booster -> Signal Generator -> Smoothing Capacitor -> Load (Bacteria)

Can I combine these values and stimulate bacteria with a pulsed 250 - 300 Volts AND have a current flowing in the range of 500 uA - 5mA?

No you can not have control of both current and voltage at the same time.

Ohms law states that :-
voltage = current * resistance
In any way you rearrange the formula you can only control one of these.
The resistance is the least controllable one because that depends on your sample and its electrical properties.
So you can give it a voltage pulse of any size but the current you get will depend on the resistance of the sample.
OR
You can give a constant current, that is a circuit where the voltage rises until a set current is reached and then the voltage is constantly adjusted ( automatically ) to keep the current at this level. Of course in this last case in practice their is an upper limit to the voltage you can supply so it might not be possible to always achieve the set current, depending on the resistance of your sample.

Grumpy_Mike:
No you can not have control of both current and voltage at the same time.

Ohms law states that :-
voltage = current * resistance
In any way you rearrange the formula you can only control one of these.
The resistance is the least controllable one because that depends on your sample and its electrical properties.
So you can give it a voltage pulse of any size but the current you get will depend on the resistance of the sample.
OR
You can give a constant current, that is a circuit where the voltage rises until a set current is reached and then the voltage is constantly adjusted ( automatically ) to keep the current at this level. Of course in this last case in practice their is an upper limit to the voltage you can supply so it might not be possible to always achieve the set current, depending on the resistance of your sample.

OK, that makes sense to me. Which is easier to control voltage, or current?

I was thinking that because current is so low in all these experiments, then in return voltage would have to be (by Ohms law) fairly high. If I chose to keep current constant at around 500 uA, and assumed the resistance of my bacteria to be a fluctuating 1000K Ohms (I have no idea how accurate this is.) then I could use a voltage regulator at around 500 Volts give or take to keep that current constant, correct?

This being done I could then (by magic) supply this output to a 555 timer to produce a pulsating square wave that I then could output on my bacterial load?

Help me if my understanding is wrong on this, but if it is correct in theory. How could I then use Arduino to program various current levels between 500uA to 5 mA?

Which is easier to control voltage, or current?

Voltage.

If I chose to keep current constant at around 500 uA, and assumed the resistance of my bacteria to be a fluctuating 1000K Ohms (I have no idea how accurate this is.) then I could use a voltage regulator at around 500 Volts give or take to keep that current constant, correct?

Correct as it stands but I would check the resistance assumption.

This being done I could then (by magic) supply this output to a 555 timer to produce a pulsating square wave that I then could output on my bacterial load?

No, 500V is a very high voltage as electronics go, most electronics work on much lower voltages. For example a 555 can only work at voltages lower than 18V. Using voltages of 500V is dangerous and you have to know what you are doing, it could kill you.

How could I then use Arduino to program various current levels between 500uA to 5 mA?

Directly you can't. You have to design a power supply that accepts low level voltage signals from the Arduino and controls your high voltage circuit. This is not an easy task, and their are some professional electronic engineers who would hesitate to take on a project like this.

This project is a lot harder than you think which is why equipment that can do this is quite expensive.

Grumpy_Mike:
Voltage.
Correct as it stands but I would check the resistance assumption.
No, 500V is a very high voltage as electronics go, most electronics work on much lower voltages. For example a 555 can only work at voltages lower than 18V. Using voltages of 500V is dangerous and you have to know what you are doing, it could kill you.
Directly you can’t. You have to design a power supply that accepts low level voltage signals from the Arduino and controls your high voltage circuit. This is not an easy task, and their are some professional electronic engineers who would hesitate to take on a project like this.

This project is a lot harder than you think which is why equipment that can do this is quite expensive.

Interesting, and I do appreciate the added concern. Rather than building a specific power supply myself, I think I have settled on purchasing a fairly cheap power supply with a controllable output of around 50 - 100 V.

With this could I then use a relay, or some other kind of switching mechanism in junction with a 555 timer, and Arduino to switch on and off my power supply to my load via controlled pulses by the timer?

And at the same time control the current of my output with series limiting resistors, such as a pot?

That way I am not directly pumping 50 - 100 Volts into the timer, but using the timers function to switch power to the load on and off?

Hi,
The resistance of your bacteria circuit will depend not just on the bacteria, but the medium that the bacteria is occupying.

If the medium has water or some relatively low conductive component then your applied voltage for any given current will be relatively low.

In the reports you have cited, what mechanical arrangement do they use to pass current through the sample?

The electrodes will also have to be inert, as an electric current will cause electrolytic action at the electrodes if the medium type/ electrode material combination is not right.
Any product of electrolysis may cause the same result you are hoping an electric current may cause, as you have cited, the electrodes caused the demised of the bacteria.

Tom... :slight_smile:
I think a long time ago I read that this method was being used in research to kill the AIDs virus.

I think I have settled on purchasing a fairly cheap power supply with a controllable output of around 50 - 100 V.

That is the nub of your problem. You have to know what sort of control that this so far mythical, power supply needs.
If their is an on / off control then yes an Arduino can turn it on and off in a PWM manner. What you need is a filter on the power supplies' output to reduce the full on / off waveform to the average voltage.

And at the same time control the current of my output with series limiting resistors, such as a pot?

  1. You don't control the current with a series resistor, you limit it. If your sample is changing resistance then I can't see this adding anything to your results, in fact it will make them more complex to interpret.

  2. A pot is not a good thing to use because at the end of the track you have an increasingly small length of track dissipating and increasingly larger amount of power. This leads to the pot burning out. An extra resistor in series with the pot will be needed to prevent this.

I think their might be some confusion amongst the researchers as to what kills bacteria or in fact any living thing. Basically it is the current. The voltage is simply a means to push lethal current through the victim.
For example it takes about 30mA to kill a person, you can get that with a 9V battery. However you must have enough voltage to push that current through the victim and that depends on a lot of factors.
Their is a little rhyme hat goes, "it's the volts that jolts but the mills that kills". A person can withstand a very high voltage 10kV can hurt but is by no means fatal, this happens in static discharge from a carpet. The resason people are not killed by carpet static is that the impedance of the static source is very high and that limits the current. If you take your 9V battery and open up the chest and apply wires each side of the heart you can kill some one, although the opening up of the chest alone would probably do it.

It could be that your bacteria are dying because the media they are in is heating up with the current and they're dying of heat, or the current is actually disturbing the functioning of the cells.

What do you want to show with these tests?

TomGeorge:
Hi,
The resistance of your bacteria circuit will depend not just on the bacteria, but the medium that the bacteria is occupying.

If the medium has water or some relatively low conductive component then your applied voltage for any given current will be relatively low.

In the reports you have cited, what mechanical arrangement do they use to pass current through the sample?

The electrodes will also have to be inert, as an electric current will cause electrolytic action at the electrodes if the medium type/ electrode material combination is not right.
Any product of electrolysis may cause the same result you are hoping an electric current may cause, as you have cited, the electrodes caused the demised of the bacteria.

Tom… :slight_smile:
I think a long time ago I read that this method was being used in research to kill the AIDs virus.

To answer your question Tom, and I appreciate the input on this topic. I have pulled a quote out of one of the papers I had received upon request from a couple of these researchers inquiring more details about their experiment setup.

Maybe you can understand this better than I can, but I do feel like they are just placing some wires into a medium broth solution (basicly salty water) with bacteria and trying to zap the bugs.

Designed by the Mayo Division of Engineering, the treatment device consisted of an 8-channel computer controlled current generator that delivers 20–2,000μA of DC; alternatively, a Keithley 2400 SourceMeter (Cleveland, OH) was used. Discs containing biofilm were exposed to DC via platinum electrodes (with the exception of initial catalase, SOD, and antioxidant supplementation assays which used stainless steel electrodes) in previously-described polycarbonate test chambers [10] (Fig 1). Electrodes measured 1.5 mm in diameter and 55 mm in length. To set up each experiment, biofilm-coated discs were removed from the well plates and rinsed by gently dipping in sterile saline to remove planktonic bacteria. The discs were then placed in test chambers in an upright position with electrodes positioned on both sides, 3 mm from the disc [20]. A 1X phosphate flow buffer was prepared with 426 mg Na2HPO4, 205 mg KH2PO4, and 640 mg glucose in one liter of distilled water, adjusted to a pH of 7.4 and filter sterilized. The stock flow buffer was diluted to 3% (30 ml stock buffer + 970 ml sterile water) for each experiment. The 3% phosphate flow buffer was continuously pumped through each test chamber at 3 ml/hour.

This how they have arranged their method of testing the experiment.

Now, to address @Grump_Mike:

Grumpy_Mike:
That is the nub of your problem. You have to know what sort of control that this so far mythical, power supply needs.
If their is an on / off control then yes an Arduino can turn it on and off in a PWM manner. What you need is a filter on the power supplies’ output to reduce the full on / off waveform to the average voltage.

  1. You don’t control the current with a series resistor, you limit it. If your sample is changing resistance then I can’t see this adding anything to your results, in fact it will make them more complex to interpret.

  2. A pot is not a good thing to use because at the end of the track you have an increasingly small length of track dissipating and increasingly larger amount of power. This leads to the pot burning out. An extra resistor in series with the pot will be needed to prevent this.

When you mention the control my power supply needs, what exactly do you mean by this? If I were to start this experiment from scratch without any prior knowledge of voltage/current levels wouldn’t I need a source of power to begin my experiment? Something I could just start with by saying, “Hey I am going to try to pump 1 A of current into this strand of bacteria and see what happens?”

I am currently trying to recreate as closely as I can using the successful parameters of these experiments to kill my own bugs, and adapt from there.

Grumpy_Mike:
I think their might be some confusion amongst the researchers as to what kills bacteria or in fact any living thing. Basically it is the current. The voltage is simply a means to push lethal current through the victim.
For example it takes about 30mA to kill a person, you can get that with a 9V battery. However you must have enough voltage to push that current through the victim and that depends on a lot of factors.
Their is a little rhyme hat goes, “it’s the volts that jolts but the mills that kills”. A person can withstand a very high voltage 10kV can hurt but is by no means fatal, this happens in static discharge from a carpet. The resason people are not killed by carpet static is that the impedance of the static source is very high and that limits the current. If you take your 9V battery and open up the chest and apply wires each side of the heart you can kill some one, although the opening up of the chest alone would probably do it.

It could be that your bacteria are dying because the media they are in is heating up with the current and they’re dying of heat, or the current is actually disturbing the functioning of the cells.

What do you want to show with these tests?

You also bring up a good point about the bacteria dying off, because of the media they are in heating up. I still am not understanding why I couldn’t switch a power supply ‘on and off’ of a set voltage across a load to maybe simulate a square wave?

If my understanding of power electronics, or power engineering is incorrect, please feel free to point me in the right direction. I am open to honest opinion and love to learn more from you guys.

Thanks again.