Do you know how to make a yeast starter? Then why not farm yeast and freeze it? - Page 22 - Home Brew Forums
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Old 10-11-2012, 02:46 AM   #211
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Are you missing any key nutrients? A little yeast energizer/nutrient may be inexpensive insurance. Something with zinc and FAN. Even for starters, I'll use nutrients.


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Old 10-11-2012, 01:22 PM   #212
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I'm using this yeast nutrient from Wyeast. http://www.rebelbrewer.com/shoppingc...52d-1.5oz.html

Their website says that's all that's needed for starters. Do you think I should be using something else?



 
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Old 10-11-2012, 02:21 PM   #213
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Quote:
Originally Posted by Forkhead View Post
I'm using this yeast nutrient from Wyeast. http://www.rebelbrewer.com/shoppingc...52d-1.5oz.html

Their website says that's all that's needed for starters. Do you think I should be using something else?
No. I think yeast nutrient is pretty much all the same.
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Old 10-11-2012, 06:52 PM   #214
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I just spoke with the microbiologist from Wyeast on the phone. Very helpful guy. He gave me some tips that I'll try to implement this weekend. He told me their pitch rate calculator is only for very rough estimates, and that actual numbers will vary a lot based on strain and equipment.

I'll post more details about his tips after I try them out.

 
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Old 10-17-2012, 08:41 PM   #215
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So my problem was a lack of oxygen in my cultures. I used an aquarium pump to do continuous aeration in addition to infusing pure oxygen at the start, and I got really nice growth.

I started with 5 billion cells in 1L and they grew up to 70 billion in 12 hrs. I then removed the supernatant and added 1.2L fresh wort and they grew up to 200 billion in 12 hrs. I removed the sup. again and added 1.4L fresh wort and they grew up to 300 billion, which is 220 million/ml. I removed the sup. again and added 1.5L fresh wort, but they only grew up to 350 billion (230 million/ml). I'm pretty sure that's the concentration limit.

In doing some research, I found the "Current protocols in Molecular Biology: Yeast" and it says stationary phase occurs around 200e6/ml, which is what I'm seeing. I don't think it's possible to get much higher cell concentration than that. So if you want to pitch at 1 million/ml/degree Plato and your OG is 1.100 (which is on the pretty high end), you'd need 500 billion cells, and a 2.5L culture to get there.

Someone correct me if I'm wrong, but I think those pitch rate calculators are incorrect at pitch rates above 200 million/ml. Also, you need to have continuous aeration if you want to get cell growth, not just fermentation of your starter.

I now have enough yeast to test different freezing conditions, which I'll do this weekend. I'll post my results next week sometime.

 
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Old 10-17-2012, 09:35 PM   #216
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Intersting. I like the continous aeration thing. Did you still use a stir plate, or did the air pump keep it agitated enough? I assume you used an air filter? What kind of stone? I think you're wrong about the calculators. They don't calculate anything over recommended pitching rates. Where are you getting 200 million cells per ml for a pitching rate? I just went to Mr. Malty, and to grow 500 billion cells with a stir plate, it recommends a 6.3 liter starter. That assumes that you are starting with 96 billion viable cells. That comes out to 80 million cells per ml if you hit your target cell count. That's no where close to maximum density. The pitching rates that it uses to determine how many cells you need are .75 million/ml/degrees plate for ales and 1.5 million for lagers.
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Old 10-17-2012, 11:52 PM   #217
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The continuous aeration idea came from the guy at Wyeast that I spoke with. I used an aquarium pump with an inline 0.2 micron filter and no diffusing stone, just blowing bubbles into the wort. I don't know if an air stone would help or not, probably not if you're using a stir plate. I did use a stir plate, and I don't think the bubbles alone would've been enough to keep the cells agitated or promote good gas exchange. The theory behind using continuous aeration goes like this...

Oxygen is required for the cells to synthesize new cell wall components during division. Thus, no oxygen, no division. You'd think the stir plate would help get new oxygen dissolved in the culture, but since the cells are making so much CO2, you get positive pressure within the culture flask that doesn't allow atmospheric gas to get into the flask. So, the only way to get oxygen in there is to pump it in and drive out the CO2. The stir plate on its own is good for driving the CO2 out of solution and keeping the cells in suspension so they don't get too crowded. When you have air being pumped in, the stir plate will also facilitate good gas exchange and keep the culture oxygenated.

I haven't explored the Mr. Malty calculator much, I was basing all of my comments on the Wyeast calculator. For example, their calculator says if you start with 3 activator packs (300 billion cells) in 1L, you'll get 483 billion total cells. I don't think there's any way you'll get 300 billion cells per L (or 300 million/ml) to expand at all let alone generate 183 billion new cells. I think it's safe to assume you'll never get more than 200-250 million cells/ml for any culture under ideal conditions. A 6.3L culture to generat 500 billion cells seems like overkill to me, but might be necessary if you don't have a stir plate or continuous aeration. If you have ideal conditions and can get to 200 million/ml, you should only need a 2.5L culture. I'll be testing that hypothesis out in the future though.

I'm using a pitching rate of 1 million/ml/degree plato for ales, not 200 million. Sorry if that wasn't clear in my earlier post.

 
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Old 10-18-2012, 05:42 AM   #218
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In my experience the maximal density you are suggesting is a good guideline for a vigorously shaken culture growing in a nutrient rich medium.
With vigorous shaking or aeration. I also think the pitching rates you are suggesting are in the correct ballpark.
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Old 10-18-2012, 04:44 PM   #219
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I see. I've always wondered about the Wyeast calculator actually. It is a quite a bit different than Mr. Malty. It's easier to use when determining step ups, which is why I've preferred it. That and I figured they knew what they're doing. I guess if you want something done right you gotta do it yourself.
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Old 10-18-2012, 05:19 PM   #220
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This calculator has fallen into favor for me.

http://yeastcalc.com/


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