Tag: fermentation

Fred Transplant: Success!

A Gram-stained view of yeasts and bacteria in a sourdough culture named 'Fred'.

I had to do a Fred Transplant last week. A grey fuzzy mold had taken up residence in on the sides of the jar above Fred’s liquid culture, so I set up a fresh container with fresh water and flour, and dipped a spoon down the center of Fred to the bottom, pulling up just a tiny amount of the stuff in there. Then I mixed it into the fresh stuff and covered it with plastic wrap (instead of a paper towel this time.)

Fred smells like Swiss Cheese Feet right now, but he’s obviously still growing, as you can see from last night’s “Gram Stain” microscopy. The slightly blurry light-red-brown lumps are, I believe, yeast cells, possibly Saccharomyces boulardii, since I dumped a capsule of supposedly-still-viable “probiotic” yeast of that species into Fred previously. I have no idea who the bacteria are in here at the moment. I did also see a small number of longer, thinner bacterial cells in there (presumably Lactobacillus) though most of them are the ones you see here.

Meanwhile, I’m about to dig out the still-unused Hillbilly Autoclave and try it out on the media I’m mixing up to try to obtain a culture of genuine wild “native flora” vinegar/kombucha yeast-and-bacteria to play with from the local wildflowers that are just now getting into full bloom.

My starting recipe goes something like this: I mix up about 2 Liters of distilled water with about 100g of glucose (“Dextrose”/”Corn Sugar”), 100g of sucrose, 500mg of L-Arginine, and enough phosphoric acid to drop the pH down to about 5.5 to 6.0. That is intended to be then poured into small “canning” jars in about 100ml amounts and pressure-cooked for at least 15 minutes to sufficiently sterilize and seal them. Meanwhile, a single generic-brand children’s chewable vitamin is crushed up and dumped into a 4-oz bottle of cheap vodka and well shaken.

Then when it comes time to go bioprospecting, I’ll pop open the jar of acidic sugar solution and add about 5ml of the cheap-vitamin-vodka to it to give me about 2% ethanol, and then go find some flowers to cut off and dump into the jars, which will be loosely covered with foil (to let air in but keep dust out) and put in a nice quiet cupboard to grow for a few days.

Hypothetically, the only things that are likely to grow in that will be microorganisms associated with vinegar-making. At some point I’ll also make up a batch of sweet black tea and see if I get a kombucha-like culture going in it, and make up some solid media to try to isolate individual microbes from it.

Amateur Soap Microbiology and my new Friend

I thought soap was supposed to be *clean*!

lumpy yellow microbial colonies growing on the soap inside of a hand-soap dispenser
People usually assume soap gets rid of funky microbes that might grow on things, so I was very amused several months ago when I spotted something growing on top of the soap in one of the household hand-soap dispensers. As of today, it looks as pictured at left. That lumpy yellow and brown mass atop the the soap looked to me like some sort of soap-sodden mold, and have been saving the dispenser specifically in the hopes that someday I’d have a microscope and could take a look at it. Meanwhile, the mass spread, and slowly started releasing some kind of yellow pigment into the soap.

Incidentally, I kind of doubt this indicates some sort of failure on the part of the manufacturer of the soap. I don’t recall for certain, but I think I may have opened the dispenser at one point to transfer some of the soap to another nearly-empty dispenser. When the mass started growing originally, it was a single spot, which suggests a single spore or speck of dust floating in and landing on the surface. Hey, it happens. Anyway, I’ve therefore blanked out the name of the manufacturer since I don’t think they really have anything to do with this.

VWR VistaVision Microscope
This mysterious growth upon my soap remained mysterious until today. Thanks to the Minister of Domestic Affairs and VWR (who managed to find me a really good deal), I finally got to actually get a close look at that lumpy mass. Meet my new friend Minnie (pictured at right). I could gaze into those eyes for hours. I couldn’t afford a darkfield condenser, and I sure as heck couldn’t afford to upgrade to phase-contrast gear, but I can add either one later if the opportunity presents itself. I also can’t afford the overpriced proprietary digital camera attachments either, though working around that is a whole other project. Until I identify an affordable model that plays well with Linux or work out how to modify a webcam into an ocular attachment,
I’ll have to settle for a trick…

It turns out if you take a digital camera and set it for close-up photos, you can actually stick the camera lens right up to the eyepiece and often get a serviceable picture.. Now, I had to subject the pictures I got today to moderately heavy processing to bring out the detail a bit better, but at least part of that is just me working on learning how to optimize the camera settings for this kind of use.

Equipped with some surplus slides and cover-slips donated by a kind professor who had some extra packages, I opened up the soap container and smeared a little of the yellow crud on a couple of them. One I just slapped a coverslip on for direct observation – the other I smeared over a slide and let dry with the intention of staining using the tiny, previously-unused vial of methylene blue left over from a very old plastic toy microscope. While the latter dried, I took a look at the wet mount hoping to finally see the mold mycelia that I had been expecting…

There wasn’t enough contrast to bother trying to get a photo, but it was obvious at 400x that what I was looking at was bacteria, not mold. Nerdly joy at learning something by looking in the microscope that I wouldn’t have otherwise known ensued, along with happiness as I realized this meant I had a perfect excuse to dig out my recent shipment from the Maker Shed – materials for doing a “Gram Stain”. Incidentally, the “Maker Shed” had the supplies on the way to me within hours of my ordering it, and they have lots and lots of cool stuff. I highly recommend it. Anyway, I got to do a “Gram Stain” for the first time in a couple of years (and the first time ever outside of a school lab). Want to see?

Mystery Microbe, I see you!

Gram-stained bacteria
Here it is – the nasty yellow goo that infected my bottle of hand-soap. My staining technique was a little off since I’m out of practice – the way I interpret the results is that what I’ve got here is neither a member of the Firmicutes (i.e. “Gram positive”) nor – probably – Actinobacteria. I really can’t guess at more than that, though. I think the few “Gram-positive”-looking cells there are artifacts of insufficient decolorization. I know I still had a surplus of the purple “Crystal Violet” stain still on the slide at the end. (How did I know? I’ll show you at the end…). The irregular bluish bits towards the bottom are, I believe, just bits of stuff from the soap itself.

Meanwhile, this pretty much satisfies my curiousity about the Mystery Soap-Infecting Microbe. There’s certainly a lot more I could investigate, but my developing Hillbilly Biotech lab is really intended to support my interest in intentional food microbiology and perhaps evenutally some small-scale non-food industrial microbiology. I have some remaining curiousity about the yellow pigment and whether or not it might be useful for something, but I’m doubting there is any food or beverage I might want to grow this stuff in and therefore don’t have much use for it. Still, I’ll keep the bottle around for a while before I throw it out in case I think of something fun to do with it. If I end up being really interested in the identity of the bug growing on it, I should be able to find a liquid that I can grow a big mess of it in, then run it through a simple DNA extraction process. Then all I need to do is find someone who can supply PCR primers, a thermocycler, and sequencing services cheap. It might sound like I’m being facetious, but I wouldn’t be surprised these days if I manage to find somewhere that’d do it for $20/sample or less. I may eventually do this will the Mystery Soap Bug anyway, since I hope to be running through this process with cultures of sourdough, yogurt, cheese, vinegar, and brewing microbes that I develop myself. For now, though, it’s just nice to be playing with microbiology equipment again. And now fully independently! Wheeeeeeee!!!!!!

Yes, I’m a nerd. And proud of it!

What’s next?

Now that I finally have a microscope, I no longer have any excuse for not getting to work on the rest of my Hillbilly Biotech lab. Just this weekend I was pricing out Hillbilly Autoclaves. I picked up a cheap air pump and air stone
for potentially building an aerobic bubble-column fermenter (for quick growth of yeast starters or a working model of a “Fring’s Acetator®”-style vinegar generator. I still want to build an ozone generator for sanitization and to get a pH meter. I’d like to also get my hands on some wheat, barley, and rye seeds to sanitize, sprout, and grow here as the first stage of developing a truly local sourdough culture, plus arrange to have several pounds of plain flour irradiated to sterilize it.

I’m also like summer to be over. Yes, I’m writing this in Winter, but it’s not until later in the summer to autumn that locally-grown fruits will start becoming available, and locally grown fruits ought to be an ideal source of local brewing and baking yeasts and bacteria. Finally, I’d like to find a wealthy patron (or matron, I’m no sexist…) who would sponsor me so I could just pursue food-microbe bioprospecting and research full-time…

Oh, yes, and I need to get around to finishing Episode 4 of my little podcast project, especially since episode 4’s topic is a fundamental microbiology technique.

Comments welcome below – thanks for reading!

Oh, and as a reward for getting all the way to the end, here’s a picture that I thought was pretty – crystals of “Crystal Violet” and iodine. I told you I had too much left on the slide…
Crystallized dye left on the slide

What shall I do?!?

Colored nigiri rice colored and shaped like 'Marshmallow Peeps'Since natural forces have not (yet) secured a replacement job for my Minister of Domestic Affairs, I’m forced to appeal to the supernatural and assume that the Spirits are angry with me for not yet having performed the appropriate Devouring Of The Soft Pink Bunnies ritual this year…so I’m devouring a package of pink “Peeps” bunnies. No doubt good fortune will finally swiftly follow…

Meanwhile, I’m poking around in patents and scientific papers, pondering a few different possible topics for the next post:

  • More on the “Nathan System” of brewing, with reference to patents and the intended purpose thereof (and pondering how to construct a homebrew-scale “Hillbilly Biotech™” version thereof)
  • Fermented foods and beverages review and request for ones I haven’t heard of yet
  • Bad poetry
  • The previously mentioned paper on getting yeast cells to eat themselves to death
  • A post composed mainly of photographs of myself with no clothes
  • The current collection of quasi-random scientific publications I’m wading through for fun
  • Some computer-related nerdity
  • Anything else

Please place your suggestions in the comments, I beg of you. Even if they are just long strings of profane threats of violence should I elect to post on [insert forbidden topic here].

“Improvements in the Fermentation and Maturation of Beers”

Judging by my webserver’s logs, almost nobody actually bothers to click through the blog-carnival host’s site to read my Giant’s Shoulders” posts. This could be due to a secret conspiracy involving famous bloggers and several shadowy government agencies. I suppose, though, that there’s a chance that simply nobody but me is that interested in non-medical microbiology. Well…today’s post is an attempt to disprove that concept, for what aspect of non-medical microbiology could be more universally appealing than beer?

Unfortunately, in the middle of trying to assemble this posting, I see the February host has decided to put the carnival up a day early, undercutting my experiment. See, I told you it was a conspiracy! I suspect the Secret Cabal of Popular Bloggers was getting pressure from the Trilateral Comission, the NSA, and Pepsico® to silence me, so they had to do it. At least being forced to miss one, I am now free from the “I’ve been posting to these since the beginning, I can’t miss one now!” treadmill.

That means, loyal readers, that you get to see this post a month before everyone else! Hooray! Stick it to The Man™! Comically paranoid rantings aside, it also means I can split this up into more than one post, which may be more readable considering how much ground the article in question actually covers. Today’s Classic Scientific Paper is:

Nathan, L:”Improvements in the fermentation and maturation of beers.”; 1930; J. Inst. Brewing; 36; pp538-550

I ran across this reference recently while working my way through an industrial microbiology text[1] that I checked out of the campus library. According to the author of this text, “The use of cylindro-conical vessels in the brewing of lager was first proposed by Nathan (1930)[…]”, referring to the now-ubiquitous style of metal fermenter seen in small brewpubs and “MegaBladderwashCo” large-scale industrial breweries alike. Based on this I had expected the reference to be a digression on the design, construction, and testing of the fermenter. When inter-library loan managed to get me a copy of the paper, I found something much more involved.

The paper is a presentation made by Dr. Leopold Nathan in 1930 to the Scottish section of the Institute of Brewing. The topic was not simply a fermenter design but the entire “Nathan System” of brewing which appears to be the basis of modern large-scale brewing, especially for Lager-type beers. At this point, Dr. Nathan had apparently already been developing this system for about thirty years (apparently starting with a German patent in 1908, which I’ve yet to find a copy of), so as you might guess it was not just a single invention but a whole collection of them. Compared to the more rustic techniques frequently in use at the time, the “Nathan System” of brewing promised to provide faster production, more consistent results, and a better final product. It does this mainly by improving the removal of “trub” (the cloudy bits of protein and such that settle out of the malt-water – the “wort” – after you boil it), preventing infection of the beer with undesirable organisms during the cooling, hops-infusion, and aeration, and by eliminating the need to “age” the brew to make it palatable. The most important improvement in the “Nathan Process” seems to be how he treats the wort between boiling and “pitching”.

—
For anyone unfamiliar with the brewing process, here’s a Grossly Oversimplified review of the steps:

  • Boil some malt-sugar dissolved in water to sterilize it and to help coagulate the “trub” proteins so they’ll settle out of the liquid.
  • Cool the malt solution and aerate it so that the yeast will grow in it.
  • “Pitch” your yeast into the now-cooled-and-aerated malt-water, in a container that will keep air out while letting out the carbon dioxide bubbles that the yeast will give of during the fermentation
  • Wait until the yeast get done fermenting, then put the resulting liquid into bottles/kegs/casks/whatever.

—

Diagram showing the containment vessel, cooling system, and sterile-air generator for the 'Nathan method' of brewing
I’ve added a couple of labels to that image from the paper, which I’m guessing was itself copied from a contemporary patent of Dr. Nathan’s. There are two purposes to this part of the Nathan Process – To cool and aerate the wort quickly without exposing it to risk of contamination, and to move trub and volatile sulfur compounds that would otherwise make the brew taste and smell funny. The hot boiled wort is pumped directly into an insulated vat (labelled “A” in the diagram) from the boiling kettle. At this stage the wort is hot enough to prevent anything from landing in it and growing. Then, the hot wort is pumped from the top of this vat into a clean-room containing a cooling device that the wort is poured on, cooling and aerating it as it flows through. Infection is prevented here by the fact that the room has a continuous stream of “sterilized” (or at least well-filtered) air, which is exhausted through the vent in the ceiling. The cooled, aerated wort is then pumped back out of the room and into the bottom of the insulated container below the still-hot wort.

Because of the large open cooling room with its constant stream of clean air, the cooling and aeration step also allows the volatile sulfurous compounds of “jungbukett” (The “Bouquet of Youth”; the unpleasant smells and tastes of immature beer, described in this paper as ‘onion-like’) to evaporate off and be carried away. Since waiting for these compounds to break down was apparently a primary reason for having to “age” lager before selling it, this not only improves the quality but eliminates the need to store the beer for months after fermentation.

The now-chilled wort then rests back in vat “A” and the trub settles out onto horizontal plates inside the vat, where it stays behind when the clarified wort is pumped out to the fermenters.

I did some poking around, and this appears to be what is described in US Patent# 1,581,194 (application filed in August of 1921), in case you are bored and want to look that up. If not, or if you don’t want to deal with the frustrating hassle of trying to view TIFF files in your browser, I intend to provide a followup post with some more details of the process and some interesting bits I found in it, and I’ll include a pdf of the patents, assuming anyone wants them.

Oh, one last thing – I’ve had no luck getting any biographical information about Dr. Leopold Nathan. Unfortunately when you search for “Leopold Nathan”, the results are clogged with references to a murdering smartass named “Nathan Leopold” instead. Doesn’t Google™ realize that brewmeisters are far more important than obscure murderers? No pictures of him, either, so I can’t even say whether his hairstyle is cooler than Eduard Buchner’s or not.

[1] Stanbury PF, Whitaker A, Hall SJ:”Principles of Fermentation Technology (2nd edition)”; 1995; Elsevier Science, Ltd; Tarrytown NY

I have a shocking confession to make.

I think I may be a nerd.

No, no, don’t try to deny it. I think it pretty much has to be true for someone who reads a 10-page scientific paper (hover or click here to see the reference) in order to learn that bacteria-snot is slimy. Yes, I am a nerd. And for that I am deeply, deeply….

Oh, who am I kidding? I like being a nerd.

Stainless Steel fermenters at a breweryFor one thing, being a nerd allows me to fully enjoy one of the perks that my job gives me – namely access to a lot of scientific papers that I otherwise wouldn’t be able to afford to obtain access to from the greedy [insert your favority profanity here] who insist on charging $30 for permission to look at a decades-old articles for a day. I should add that this perk includes Inter-Library Loan for articles that I can’t get online, and the service on campus is great so far. Same day delivery of a classic article from 1930 in what I’m guessing most people would probably consider an obscure journal.

It doesn’t have quite the same thing in it that I expected from the source that pointed me to it, but I think it can still be considered “classic”. I need to re-read it more carefully to make a final decision on this, but I think I have my next “The Giant’s Shoulders” article in time for this month’s upcoming issue. And, yes, the picture attached to this post is a hint (and, no, it’s not directly related to the bacteria-snot article mentioned above in any way…)

This Week = No Fun, but here’s an update anyway…

Busy week with unpleasant surprises, but I ain’t dead yet. You’re probably wondering what a hot dog that’s apparently eagerly anticipating eating itself has to do with that. The answer is: nothing, but it does relate to something I have been intending to post about for a long time, but haven’t since I didn’t have access to the paper…

So, in lieu of blasting out Twitter®-style updates on my Laconica feed that nobody reads anyway (a few people no doubt see the echo of them on Twitter® itself, but I don’t know if anyone cares…), here are a couple of what-I’m-doing-now updates before I go to bed:

  • I just shot off an email to the webmaster (the only contact I could find who might have the relevant information) of the Institute of Brewing and Distilling, trying to get access to a classic paper published in the Journal of the Institute of Brewing and Distilling for the upcoming “On the Shoulders of Giants” carnival.

    • There’s no way I can afford the ~$300US that it would cost to join the group, nor even at the moment to pay the typical $30 or so that greedy paywall-imprisoned publishers charge for individual articles. However, they appear to be opening up their archives to the public, for which I think they deserve substantial praise. Still, they haven’t worked their way back to the first few decades of the 20th century yet, so I had to email and ask if there was a way to get the article in question. If not, I’ll see if there’s any way to get it through inter-library loan in time.

  • “Small Things Considered” asks “What Microbiological Discovery in 2008 Did You Find Especially Interesting?” Which brings us to the “self-eating” thing.

    • The paper on caffeine’s induction of macroautophagy (“self-eating”) in yeast (and what happens when benzoic acid is also present) finally escaped from the paywall prison in January, and I now have a copy. I at least thought it was full of interesting implications (along with some useful knowledge), so I’ll hopefully be posting about it soon.

  • I’m also a computer nerd

    • Particularly when it comes to things like Linux. I’ve been thinking of trying to do some recordings on a couple of practical subjects that interest me for “Hacker Public Radio”

  • And of course, I still need to do some Xanthomonas snot Xanthan Gum experiments.

I am an attention-whorewilling to listen to my readers’ interests, so if anyone has suggestions or comments feel free to post them…

Coming Soon to a Bigroom.org blog post near you!

I got a little money for Christmas, but I’m feeling quite guilty as I didn’t really have the time and money to reciprocate in advance. I also haven’t been able to figure out what to spend it on until now. Now, I have a solution for both problems.

Xanthomonas campestris growing in a petri dish and exuding slimeI intend to spend it all on fermentation-related food ingredients and do some experimentation with sweet-tasting foods. Initially, in addition to flavors (spices and whatnot), I need to track down bulk quantities of:

  • Erythritol, which is a virtually non-caloric sugar alcohol which unlike sorbitol and so forth is not normally prone to cause gastric distress, and unlike Xylitol is not hazardous for beloved household pets. Better still, it actually is very tasty unlike that nasty hippy “Stevia” crap (which isn’t produced by fermentation anyway, as far as I know).
  • Food-grade Glycerol (“Glycerine”), which I hypothesize is close enough to the structure of Erythritol to mix well with it and help the erythritol dissolve (and hopefully prevent crystallization, much the same way the “corn syrup” does with sucrose).
  • Xanthan Gum. MMmmmmm…edible bacteria-snot. (Okay, for all that this sounds disgusting, it’s really somewhat similar to pectin, which like xanthan gum is a polysaccharide. Pectin is just fruit-snot rather than bacteria snot. Dietarily, both count as “soluble fiber”.)

There are probably other ingredients I can come up with as well. For those of you out there who are owed gifts: Chewy candies, hard candies, baked goods, and/or beverages, what’s your preference for my initial experimentation? Assuming anyone’s interested, I will probably blog my results…

Meanwhile, I’ve also been thinking about geolocation, geotagging of audio and video media, and Asterisk again. I want to take the lessons learned from my playing with the “Where Was I?” prototype and turn it into a real geolocation system, integrating with Asterisk and Laconica (which turned out to be easier to set up than I’d feared – I’ve now got my own Laconica server at http://omniblab.dogphilosophy.net/laconica, though I need to sit down and activate the IM integration (Twitter may have abandoned IM, but it reportedly works fine in Laconica).

It also turns out that you can use Asterisk with cellphones(!), at least if they have bluetooth. That’s handy to know…

Argh – too much to do, not enough time!…

Über alkoholische Gärung ohne Hefezellen

In my last submission to “The Giants’ Shoulders” blog carnival, we saw how the famous surgeon Dr. Joseph Lister deftly demonstrated definitively that fermentation processes were caused by live microbes rather than some sort of mysterious soluble substance that just happened to be associated with microbes. In today’s episode, we will see how Eduard Buchner definitely demonstrated that fermentation was caused by a soluble substance that was associated with microbes, and no microbes are actually needed.

Better still…they’re both right. “Wait…what?” Read on, O Seeker of Microbiological Knowledge, and be enlightened by this month’s entry: Continue reading Über alkoholische Gärung ohne Hefezellen

Nerd Reading Spasm!

Did I mention the place I work has some amazingly spiffy perks for a nerd like me?

Last night, I was poking around pubmed looking for references to yeast and erythritol (namely, do yeast interact with it, and will they metabolize it?) I found precisely one relevant reference. From 1975. In a Czechoslavokian microbiology journal. A no-longer-existent Czechoslovakian microbiology journal. Even though it was a journal published in English, I didn’t figure I’d be able to find the article I was looking for. It did turn out that the greedy (insert long string of profanity here) anti-open-access “SpringerLink®” Netherlands organization has an electronic copy of the article…which I can get limited access to for a short time for a mere $34.00. Not going to happen, obviously.

Just in case the college had a subscription that would let me get to the article at no extra cost, I checked. No such luck. But…

…The campus medical science library just two buildings over from where I work has dead-tree editions of essentially the entire journal! Im name des Nudelmonster! Instead of paying $34.00, I got a photocopy of the article for about $0.50. Bonus: As I had hoped, the article[1] reports that erythritol is not metabolized by yeasts, although it is taken up to a small extent. That means I can add erythritol (or xylitol or sorbitol or whatever) to must or wort, and it’ll still be there when the yeast finish, leaving the resulting beverage still sweet. Hooray!

Plus, I was also able to get access to an electronic copy of a review of the uses of poly-?-glutamate[2], which I was bemoaning not having access to over on an interesting Small Things Considered post recently.

Speaking of reading, one thing I really could use are any worthwhile books on the general subject of applied/industrial microbiology, bioprocess engineering, fermentation, and so on. “Worthwhile” here means practical texts that are A)primarily about microbiological processes (as opposed to, say, bioengineering of plants) B)Reasonably technical, and C)Either “not very old” or “very old indeed” (I collect old science books).

I’m not a fan of Amazon.com’s abuses of the patent system, but I’m in a hurry since it’s past my bedtime already. Therefore, purely as a sampling of the kinds of books that sounded interesting to me, here is a selection in more or less random order of books that came up in a quick search on amazon.com. Anybody out there have any other suggestions?

Continue reading Nerd Reading Spasm!

Aqua-pedestrianism and Ice Cream Yeast

A teaser image from the interactive map below...Today’s batch of blog-based Stir-Fried Random includes another interactive map of a lake-spanking expedition, a very brief musing on search engines, and a return to “intentional food microbiology” discussion. To preview: you can get pizza without ever getting out of the water on Lake Conroe, “spanking” is amusingly popular for search engines, and no, there is not normally any yeast in ice cream, but perhaps there could be. Read on, please…

Continue reading Aqua-pedestrianism and Ice Cream Yeast