Month: May 2021

Ice water hash - a complete guide from harvest to dab

The amazing title picture was taken by the one and only Even Stone. Follow him on Instagram!

What is water-hash?

As you probably already expected from the heading for this method of separating trichomes from cannabis flower, we use a combination of ice and water. This mechanical procedure uses the force of the water vortex to brush of the oleoresin containing trichomeheads from yours buds. The ice in ice-water-hash making is only used to cool the whole mix down in order to make the stalks of the trichomes brittle so that they break of more easily. But the exact steps you need to take and much more will be discussed in the following article.

Why choose water-hash?

When you hear about ice-water-hash most times it sounds pretty labourintensive and it is, but the ratio of quality per worktime is the highest of the different solventless methods. Dry-ice-sifting for example oxidises the trichomes so fast that they burst and leaks out the desired compounds like the really volatile monoterpenes (limonene). And not only the trichomes burst, the plant material gets also really brittle, really fast so the contamination rate also increases rapidly. This results in a drastic loss of quality and worth, because even when pressed this can only achieve low prices on the market.

Then why not use drysift, there's no dry ice used and no enhanced oxidation. Thats completely true, but as we're going for most quality per time drysifting also loses in that category for it's time intensive cleaning process of the material to get to 90% purity. Don't get me wrong I love drysifting and such fire as this fullmelt drysift from CubanGrower x The Cuban Hash Queen (Pheno by TheVillage/Photographed by Erik Nugshots/Bred by Symbiotic-genetics)makes it worth it.

But to be profitable with that kind of premium product is really difficult and the market is not that big. So in order to sell at a lower price, but still nearly the same high quality as with drysift only ice-water-hash remains an option. "Why not just blast it all"?

Good question, and yes you could do that, but the price for bho currently is not as high as decent hash rosin. Additionally the startingcost is really high if you want to do it right. If you need deeper info on hydrocarbonextraction and it's specifics, visit Murphy Murris Instagram account

And for home users I wouldn't recommend it either if you don't have a proper, safe setup and are quite experienced with hydrocarbon handling. Hash on the other hand doesn't need a C1D1 room or other security measurements than a rubber mat on the slippery floor.

So if you want to make fullmelt the fastest way possible use ice-water-hash to separate your resin like the italien hashmaker slite23 from Barcelona with this beautiful Sundae Driver 149/90u Fullmelt (Pheno by Alorganics). Now we will look a bit further into the mechanics behind hash-making in order to build a good knowledge base, that will let you achieve resin like this with a bit of practice.

The principles of Hash-making

Terminology

In the first part of this chapter we will look at some basic terminology that we need to describe the process precisely

• Micron: This term refers to the opening space in the bubble bags. It tells you how many micrometers the wholes in the screen are wide. These mostly get abbreviated as "u" or "μ". To put that into perspective, a human hair is 75 micrometers thick and our eyes can only see down to 40 microns. Thats why it's important to buy a scope and look at the holes in the screen to see if they are all open and not clogged with nylon or bad stitching. Always wash your bag properly before using them. For washing the first time after buying new bags, I would recommend to make a 10% alcohol/water solution and rinse the heavily. But be careful not to rip the seams.
• Terpenes: These pure hydrocarbons are a part of the famous entourage-effect firstly mentioned by Dr. Ethan Russo in his paper "Taming THC: potential cannabis synergy and phytocannabinoid-terpenoid entourage effects" (see source 1) They induce a synergistic effect in combination with Cannabinoids in your brain, basically pushing the high in a certain direction. Like Limonene for example, that on its own already causes a sort of high if you breath it in at a certain concentration. Similar to that strains with high linalool make you relaxed and the same for other strains with a lot of terpenes. There are also other components like thiols and alcohol esters in Skunk e.g but they aren't as well researched as terpenes At last there are also terpinoids, which are similar to terpenes in effect and structure, but the have additional chemical groups. But because of their similarities we mostly count them to the terpene content.

• Fullmelt: We already used this term in this article and some of you probably wondered what that means. It stands for the quality of the hash in regards of melt % tage. Hash that melts like water in your banger and nearly leaves no residue is the best of the best in hash. At room temperature it sometimes already melts into a oil like mass of trichomes. More info on what differentiates the melt further and how you classify it will be discussed in the chapter Rating-Systems
• Contaminants: All particles that aren't trichome heads be it hairs, skin, dust or plant particles are considered contaminants. More on how to prevent them and proper hygiene in the coldroom in the designated chapter
• Cultivars: Every plant that is derived from sexual reproduction is, like humans, different even from his siblings. This term refers to one unique genotype with a specific phenotype that gets expressed. So for example just because some Cake clones yield not all of them will and even if you have a selected cut it might not dump a lot of weight if grown under completely different conditions like those that it was selected in. That's why it's good to ask the clone seller how it was selected or select your own cultivars
• Flowrate: Like in coffee brewing you can control some parameters like speed of water drainage and with that force that pushes through the screens. Using that you can safe time by not having to spray and rinse that much. But that needs the perfect speed and that takes practice as you have to be careful not to blast the water through as that could drag contaminants through the higher micron screen and also degrade your trichomes by removing more terpenes than necessary. But not enough pressure prevents the contaminants from getting flushed through
• RO-Water: Growers already know the term, but for everyone else: it refers to water filtered through a reverse osmosis filter. This device pulls out all the minerals and other compounds that could affect the taste. For optimal results you should make your own RO-Ice cubes (Protip: Insulate your washing vessel to save ice and prep time)

The Separation Process

How exactly can you isolate the trichomes without plucking them off one by one? For that to work we need Physics. Trichomes are secretion-organs with bulbous, oleo-resin filled heads on them. The oleo-resin is trapped by a layer of lipids that releases terpenes and other compounds slowly over the day on the plant to fight of pest. It also shields the flower from UV-Damage so to increase your trichome-coverage you should incorporate UV in your lighting mixture.

As we know from chemistry class, fatty substances don't mix well with water as they're non polar. This phenomenon is the reason why trichomes stay intact while beeing washed most of the time. If your strain as a to thin lipidlayer the trichomes smear in the bag and you won't catch most of them

That's why some cultivars won't produce hash yield numbers that justify washing the material. Even if it has a lot of good trichomes you have to separate them completely in order to catch all. Therefore they have to be evenly brittle, but not brittle.

Thats where water comes into play. This natural "solvent" stays, even with ice, at 0-5°C for the whole washing-process to insure maximum Quality and efficiency. Now that we have the Trichomes ready for action we will start the stirring process. If you're hand-washing try to get a small water vortex as the current of the water strips the flower from the Trichome heads.

If you picked a good cultivar the water should turn slightly golden/purple (depends on the strain) and you should start to see the heads drop to the bottom of the wash container. If you think you got most of them, you can stop agitating and let the trichomes settle down. After that you skim the water on top that doesn't contain resin and either filter it or throw it away. Let about 1-2/3s in the wash vessel and strain them through the Bubblebags.

How to collect the Resin from Water

But in which order do I have to put the bags in? We go from big to small openings but how many bags you have depends on your bag-set. Different companies also use different mesh sizes but most of them are in the same region so we want to give you a small List of bags that there are.

• 220μ = Filter-/Workbag: most of what's catched here is dust and plant material. This is to clean up most of the contaminants before catching anything. If you buy one that is big enough to fit into your washing vessel, you can wash in it and pull out most of the plant material even before emptying into the filter vessel.
• 180μ = Second Filter: Most bag-sets don't have them but they're a good option if you want to add another filtration process to your regimen. Most of the heads catched here also don't have a lot of value
• 160μ = Rosin-/Foodgrade: This bag collection is worth it, especially when you grow strains with really large trichomes like a GMO. You can even find some partial melt in here, but I would recommend to turn this into rosin or edibles/RSO as most of the time it is still a bit dirty.
• 120μ = great Quality Resin: This is where the real fun begins. In this range you start to see real fullmelt or at least 4 Star quality, that is either worth keeping as hash or if it's doesn't melt fully to press it and sell it for premium prices
• 90μ = exceptional Melt: The melt coming out of this bag is the true Creme de la Creme and goes over the counter for astronomical prices up to 130$/g. The fullmelt nearly leaves no residue on the banger and the taste is amazing. As said before, all non melt result should be processed to rosin or oldschool hash (Frenchy Cannoli-style)
• 70μ = is used more like a substitute for the 90μ bag. To test which one you like better, you should test wash and compare how the cultivar performs with the different bags. Most intact, ripe Trichomes are not smaller than 70μ so this will be the last bag with exceptional results.
• 45μ- 25μ = Foodgrade: This bag catches some debris, but mostly unripe or already dried up trichomes so it can be used similar to the 160μ
• "Full-Spectrum": This term is a tricky one, because a real full-spectrum Oil refers to it containing the whole range of cannabinoids and terpenes. But most Water-Hash fullspec is a 45μ through a 160μ or similar micron sizes. This leaves out a lot of other compounds so technically it's not a fullspectrum oil in my opinion. But as with everything I teach, I would be really happy to discuss these topics in the comments. Real fullspec is only achievable with solvent-extraction as it can extract everything contained in the flowers.

 

What do I do with the wet Hash now?

Now that you catched all your trichomes, you want to make them smokable by drying them really thoroughly to prevent it from molding. This can happen in the dry stage aswell so be fast with the process. There are two main options for drying Ice-water-hash. The first and cheapest option is air drying it by freezing it immediately after collection and spreading it out really really fine after that. How to do this and the exact steps will be mentioned in the drying chapter.

The most used method today is the use of a lyophilisation ofen also called freeze dryer. This neat piece of equipment uses certain physical phenomenons to quickly remove the moisture from the trichomemass. It also is the most terpene and colour preserving technic for drying. For commercial ops this device is essential for production speed and success as a failed air drying attempt could cost you a lot of money.

How to prepare yourself before setting up a hash lab

Now most of you probably would want to immediately wash their whole crop, but before doing that, we need to set up a proper lab and everything that goes with it. For starter we need the most essential item to be of the highest quality possible. Of course I'm talking about the starting-material as it is our base for good hash. That's why the mantra for most hash makers is: "Fire in, Fire out" To give you an idea of what I mean with that, we will look at our material in depth and disect what makes good Hash starting-material

The different aspects of good startingmaterial

Live vs cured Flower

The first choice with have to make after harvest is whether we want to make "live" or "cured" hash . These terms refer to the drying process of our material. Live Hash is taking the fresh flowers and directly freeze them for washing. That way, you can conserve the most terpenes especially those that flair of easily like monoterpenes. This brings more sharp, intense flavor that many people nowadays really enjoy. also the colour gets "better" more easily with this method as nearly no degradation happened. What we mean with better and which role colour plays in hash, can be looked up in the designated chapter With this method it's vital to have an extremely clean freeze, if possible you should have a separate one only for fresh frozen flower. But if you only wash once every 2-3 Months this isn't really worth it, so you can use tupperware to protect your buds from the other freezer content.

Often the whole plant gets frozen for this purpose so you will sometimes see the term WPFF on your bought hash. This stands for "Whole plant, fresh frozen"

Cured Resin on the other hand has a more "rounded" flavor as the highly volatile Monoterpenes have flaired of or turned into another terpene profile. Most of this is still in research, but you definitely taste a difference. This rosin is bit harder to get as light in colour as the live version, because the dry time has to be perfect. But as we will see in the further chapters, colour doesn't always say much about quality.

How to visually rate Material

Now that you picked your methode of choice, we can start to analyse if our product will be cut out for making hash. As mentioned before the trichomeheads and their size play a mayor role in it.

First of all lets look at this depiction of a trichome. This shape is what you're looking for in the perfect trichome category. It has a large bulbous head with a really thin "neck". This is what we want as it will make it easier for us to break them off and separate from the plant material. What we don't want are secretory glands without heads, their "hairs" may look frosty af on a plant, but if you wash them, you will be really disappointed. A typical strain with these attribute is purple punch. It looks fantastic but washes really bad most of the time. Another unfavourable trait for hash making are varieties with really long trichome stalks, small heads and a thick waxy cuticle as they won't contain a lot of active compounds. These strains are also valuable, but just not for water hash.

Don't get me wrong, if you select a fire cut, it can dump, but you would have to go through a lot of cultivars to find one. I recommend 20-50 regular seeds depending on the breeder. You can do smaller hunts, but the probability of finding a keeper cultivar that satisfies both quality and quantity aspects will be quite low. 

 

 

 

 

 

 

 

 

The importance of hash specific breeders

This is where the breeders come in. If you want to select a hash-cut but don't have the time to sift through hundrets of plants, you should pick a breeder that selects for hash. That will give you a lot higher starting chance of finding a good specimen as the breeder has already pre selected the genetic material. There are lot of different great breeders and most probably I will forget to name some and I'm sorry if I left someone out. But to give you a direction, here are some breeder recommendation from my side: Karma genetics, Oni seedco, Bloomseed co, Dying breed seeds, Archive Genetics, Truecannabliss and Cannarado to name just a few.

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Some strains especially are good for hash-making as they have a high rate of successful cultivars. Names like GMO, Sour diesel crosses or the famous Gorilla glue are well known for their ability to dump wpff yields upwards of 5%. Even most crosses with there lineages will give you your desired results so they're a good starting point for your hash-journey. For companies I would also recommend to implement a really heavy yielding strain at all times in their rotation as a safety net, if some new selections perform worse than expected.

Now that we have picked a good strain, we can start growing and watching it. When the heads are nearly finished you can scope it with a jewelers lupe or a electronic microscope. This should be done extremely thoroughly because if you don't do it and do a test wash, you potentially waste money and thats not good when you're starting a company. To give you a comparison chart we depicted one picture from a study by Dr. Potter

Figure D depicts the best usable trichomes as we don't need excessive force for separating these from the stalk. Trichomes like in figure C aren't the optimal, sure they have a large head and contain more oleo-resin than the ones in figure B, but they require so much force to get separated that there also is a lot of potential for contaminants to get into the product. All these criteria are only the beginning of a long r&d (research and development) journey that should look at terpene-make-up, terpene Content, rosin yield, stability, colour etc

The distribution of Trichome size

Another important performance indicator is how much a strain dumps in each quality area. A cultivar that puts nearly all its resin out as fullmelt is much more valuable than one that produces 70% edible grade hash. As with every property of hash that we look at, we need to do our tests here as well.

For this purpose you should use a register/excel sheet to get an overview of your data. This data is a treasure chest of usefull information about your process, that can give you the opportunity to optimize every little detail. This would be a example of how to analyse a strain:

• Terpene make up
• Terpene content
• Yield WPFF vs Cured
• Market value
• How long is the overturn time (harvest-harvest cycle)
• Trichome Size

How to test your cultivar for yield beforehand

To test and find the perfect cultivar is quite a challenge so I highly recommend keeping a cut from said plant. Not only for hash making, but for sharing /selling the clone. The high end concentrate marktet is really hot right now and new, good clones go for a lot of money. So this would be the perfect example for horizontal expansion of your production line.

But to get such a cut, we have to select and before selecting we should do some tests. The first is a cannabinoid and terpene test which you need regardless as a commercial cultivator. If you buy flowers you should ask for their Coa so that you can compare your yield to the given data and check if you catched all the trichomes. This is also a great opportunity to fine tune your normal washing processes. So if you don't get the same % in yield that is given in the testresults you should go through your process step-by-step in order to find the mistake. If you don't find anything, it could be the strain. Look at how the resin handles the water and if it smears the bags. Most times this is a large opportunity for product loss.

If thats not the problem, try using the leftover material for edibles/rso, that way you can use every little bit of the material and minimize loses.

Another test is the mason jar shake. This simple method has a bit more to do with experience, but is also usable for beginners. You simply take 10-15 grams of your desired flower, put it in a mason jar together with Ice and water. After letting it sit for 2-3minutes you can swirl it around a bit and then you should see golden trichomes dyeing the water golden. Let it sit for another 2 minutes without stirring and look at the bottom of the jar. There should be a small layer of trichomes if you have a fitting cultivar.

Cultivation Techniques for better Hash

The best way to get great hash yielding flowers is to grow them yourself, so that you can learn the subtleties that go into growing the best hash flowers. One sure way to increase resin content is adding UV-lighting to your grow as we mentioned before. This lets the flower build up its defences against this high energy radiation. Not directly for resin content but for terps, most hash cultivators use organic living soil system to grow as this lets the plant fully express its genetic potential. This debate is highly discussed in the hash community so I don't want to get too deep into it. But it has to be said, that salt based fertilizer can also achieve great hash if they're perfectly tuned in for the cultivar. A general booster for terps and secondary metabolites is a healthy amount of sulfur in our regimen. Organic growers can use gypsum or epsom salt for this purpose

The necessary Equipment for Washing

Now that we know our basics and how to choose material, we need to look at our setup. To configurate it the best way possible we need to choose between the two kinds of washing. A pros and cons list should be sufficient to give you an idea of what you want to choose

Hand vs Machine Washing

Pros washingmaschine

• Multiple maschines can be operated by one employe
• Less manual labour
• simple SOPs can be made for unexperienced operators
• "easy" for beginners

Cons washingmaschine

• Lots of work to keep perfectly clean
• High initial investment cost
• cheap maschines need a draintube change in the beginning
• often not very customizable

Pros Handwash

• Easy to clean vessels
• Fully customizable washing program
• Really gentle on the material
• Smaller initial investment
• Large batches of 10-15k grams possible

Cons Handwash

• hard physical work
• Not automated
• Experience is needed to get perfect results

The next big decision is whether to air dry or freeze dry. But it would blow up this chapter so we will look at this in the specific chapter.

A short Equipmentlist

Now for a better overview we collected a list of small and big items every wash room needs in order to function properly.

• Bubblebag-Set (+ Workbag)
• RO-Water
• Ice-cubes (Ro-cubes if possible)
• Washing Device (Maschine vs Vessel)
• Stiring device for handwash (stainless steel paddle)
• Drainvessel for filtering the hashwater (should be big enough to mount the bags in it)
• Thermometer for water temps
• Extremly clean work enviroment
• AC for cooling the room
• Cleaningagent (isopropanol 70% works best)
• Waterhose with RO-water to spray down the bags
• Cold Spoon for collection
• Dryingdevice (Freeze Dryer or Microplane/Sifter)

How do I prepare my tools?

The most important factor is cleanliness, I know I sound like a broken record, but as everything gets concentrated with hash making, the dirt also gets concentrated. Thats why the phrase: "cleanliness is next to godliness" fits perfectly for hash making. You should start with your wash room and wipe everything down with a 50/50 alcohol/water mix. Because if you only clean your tools and put them back into the room, they get contaminated aswell. Even the ceiling and the walls should be cleaned as dust can settle there.

Next we need to take a look at our tools. They also should get wiped down and put in a cold place as the resin won't stick as much then. Your Bags should also get washed with a very mild alcohol/water mix and then rinsed with RO-water. After that you can put them in the drainvessel over night in order to dry them.

If you make your own ice you can make it the night before with RO. The best shape in my opinion is half round about the size of your thumb. This minimizes the surface and thus lets your ice stay longer like this.

How to keep proper hygiene in the coldroom

We explained how to clean your room + tools, now we go to the steps you can take yourself. The biggest vector of contamination is the human, so in order to mitigate this risk we wear the corresponding PPE (personal protection equipment) Lab coats are a standard in most hashlabs but a full-body tyvec-suit would be more appropriate for this usage as it contains the dirt on your trousers as well.

As you can see in the picture a mask and glasses are also a good thing to contain bacteria in your breath and skin pieces. But thats more important if you're in the medical sector. For home use you can get away with a suit, a hairnet and proper nitril gloves. These are really practical as your hands would get sticky really really quick without them

A coldroom buildout

Now that we protected ourself and the hash from contaminants we need to build a coldroom that makes our job as glorified janitors easier.

The best working surfaces

The first thing we should direct our attention to are the surfaces. Every desk, floor and wall should be wipeable as they collect dust. The best option would be to use stainless steel everywhere because it can be safely sterilized heavily without showing any degradation. Another reason is the faster cooling off the room because of it being metal. But such coldrooms are really expensive and only for professional operators For the homeuser, I would advice to lay out paintersfoil on the floor and walls. Looks like out of the movie american psycho, but trust me it is worth it. Also it is quickly removed if some uninvited guests want to look into your flat

I would also clean the ceiling with a broom or a mob and some water if possible before washing. If you want to be really thorough, you could put in an air filter. You can use a normal carbonfilter similar to the ones in growtents.

How a professionally built shell for your coldroom

https://youtu.be/zBCXAs3xFVY

Optimal climate control

For washing and separating we need constant temperatures around 0-10°C to get the best results. Otherwise the hash will get greasy and really bad to collect. This is quite difficult if you don't have well insulated place to wash. That's why we recommend using cellars or insulated sheds for home users. If your growroom is clean and sufficient you can use it as well, but every room needs a strong AC to wash in the summer months. But how do we know which one to buy? This depends on your budget and coldroom-size. Most Acs are measured in BTU/H or Watt/H which can bei converted into each other. 1000BTU/H are equivalent to about 300 watts/h and how many of those you need can be looked up in the chart we made

Space	Watts
Up to 30m2	2350
30-37,5m2	2500
37,5-42,5m2	3000
42,5-50m2	3500
50-60m2	4000

But not everyone can afford the best Ac so we're going to look at some models for each price class

Small to medium budget

In this category we have the well known mini splits, which most of you will have in their grow room. This modell is really efficient and in my opinion the best choice for small to mid sized ops as you can get down to freezing temperatures in a medium insulated place because it doesn't use air, but a refrigerant. They're powerefficient and don't take up to much space, but it needs a second device on the outside of your building which has to be connected via copper wiring. That and the cleaningafford are the only negativ points about this system. You can get them as cheap as 800$, but you have to count in the installationcost if you're not doing it yourself. But DIY is only recommend if you're a professional or have done it before as improper installation could be a fire hazard. 

Central AC Units

These ACs are really powerful and normally used to cool entire houses. Thats why they can be quite expensive and use lots of power. If you buy one of these and use them all the time, you should think about buying a solar panel as this would pay for itself pretty quickly. This model is mostly used when already preinstalled in a large grow or a housingcomplex You will need a licensed electrician for this installation as this is really complicated and easily done wrong, which again poses a fire risk

How to setup a coldroom for good workflow

Now that we set up our basic shell for the coldroom we need to bring in our equipment. If we want proper workflow for e.g. commercial use (most hash/time) we have to think strategically on how to setup everything in an order where you don't block others from doing their part. So in order to give you an idea what one could look like, we depicted a example in here

If you don't understand all the terms, no problem, we will now take a look at the step by step tutorial. This should clear up most of the confusion

A step-by-step approach to washing Ice-water-Hash

So we have our room prepared, material sourced and ourselves clothed accordingly. Let's beginn the best part, the wash It's advisable to really check if you have everything you need in the room as opening it again could introduce more contaminants in your room Have the icecubes stored in the freezer aswell so that you can directly take more if you need it

Our material should either be frozen or dried and ready. First we prepare the material by breaking down larger nugs in thumb sized pieces in order to get the optimal surface area while not making them so small that they get grinded up between the ice. Be really gentle with the buds as every handling "destroys" trichomes and with that, reduces your yield.

When this step is finished, we continue to loading up the wash vessel. First put in the workbag, then a thick layer of ice to cool right off and keep the material from being scrubbed against the bottom of the vessel as we're trying to not damage the buds too much. Now put in a layer of material in (a bit thinner) and then a thin layer of ice again. Repeat this till all your material is in

After this, it's time to get it soaked. Use a gentle setting on your water hose and distribute it evenly over the mix until everything is floating. You want the right ratio of ice/water in order to keep the mix cold but not grind the material between ice cubes. Again, the water vortex is whats separating the trichomes from the flower. The best indicator is listening when you stir it up. Maybe do a testrun beforehand without material to get the mix right. There shouldn't be any crushing noises instead only a light collision sound of single ice cubes hitting the side of your vessel. For a example of this sound and maybe one of the greatest videotutorials out there visit Frenchy Cannolis Youtube channel for his course on hash

Let the material sit for 5-7min for WPFF and 10-12min for dried flower in the water to resoak. We do this to let the flower get more flexible so that it doesn't release to many contaminants when agitated.

Then we beginn with the stirring process. Remember to be gentle to the material, but stirr it thoroughly to get everything nicely mixed. You can start with clockwise stirring or a canoeing like move. Continue this for 5-10minutes depending on how golden the water is and how many washes you want to do. If you want to get everything out in one wash and don't care too much about getting the cleanest of the clean fullmelt you can go long 10-15min. This method is quite time saving if you only go for rosin. If you want to separate every grade of hash you go for 5-10min depending on the strain/stiring strength.

We stop stirring and let the trichomes settle for 2-3 min again so that we can skim of water from the top. We do this as it would be really heavy to empty out the vessel in the drainvessel if it's completely filled. If you're running a maschine you only set the timer and come back to open the drainvalve on the bottom to release the water into the drainvessel. I would advise you to take your sprayer/hose and spray down the drained leftovers again as a lot of hash gets stuck between cubes/material if you only drain it from the bottom. Now that we have our bags "filled" we spray down every bag down like this to get contaminants and contaminants pushed through. We continue this with every bags because trichomes that are the right size don't fall through if they're in the right bag (micron = trichome-diameter) and the contaminants fall through to the last bags if you did it right. How to that properly we will watch a video of eldaggy of him spraying down his hash

https://www.instagram.com/p/B-P_mojBYwE/?utm_source=ig_web_copy_link

Afterwards we can scoop the hash from the bags with our spoon that we put in the freezer earlier. It's advisable to cool 4-5 spoons as they warm up pretty quick again. Now depending on our dry method we put the hash on our trays and spread it evenly to not get any water pockets. If we freeze dry we can put it aside/directly in the dryer. If we air dry we need to squeeze out most of the liquid with a 25micron screen, cover the hash and put it in the freezer in order to microplane/sieve it later on. We can repeat these steps until no more hash comes out or we don't like the work/quality ratio anymore and use the rest for blasting/alc extraktion

How to dry wet Hash

As we can't smoke wet hash we have to dry it now. And as said before we need to decide between air/freeze dry as a home hash maker. For commercial washing you can't use airdry as the risk of productloss and the dry time will make it unprofitable. I mean you can do it, but in order to make enough money of that you need brandawarenes.

Whats a Freeze Dryer

This lovely device is also called a lyophilization ofen and dries your hash in a short time. It does this by freezing it down to about -40°C and then slightly heating the trays with the hash, causing the remaining moisture to directly sublimate into a gas. This effect is encouraged by a vacuum that you pull on the drying chamber. This leaves the highest possible content of terpenes in your hash and prevents it from oxidizing which would darken your hash aswell. For info on how exactly this works and how a freeze dryer looks like we will watch a short video from a big drying manufacturer

https://www.youtube.com/watch?v=eE993Vs7j8s

For the tech heads under you, we have a second option to buying a prebuilt unit. There is the possibility of building your own freeze dryer. This will safe you a lot of money, but beware, it won't give you the same results that a commercial, specialized unit will give you. Here's a short video tutorial

https://www.youtube.com/watch?v=obhYSf1ogoo

How do I choose and use a Freeze Dryer

Let's assume you want to buy one of these babies but don't know where to look. We would recommend different models depending on your capacity needs and budget. First we have the harvest right models, these are what most people use in the industry as they beginn at 2500€ and a capacity of 800gs of Hash each day of usage. The only problem is the customer service and life time of these maschine as they require intensive care especially with an Oil pump. The second brand are labconco dryers. They get used in the pharma space and are the Ferrari under freeze dryers. These units are built for a lifetime, but also really expensive as they begin at around 15000€. We would recommend them only for commercial users as they won't make sense for the home user.

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But then there is another decision to make. Do I go for the cheaper oil vacuum pump or do I invest in an oilless version

This mostly depends on your budget as I would advise you to always go with the oilless version as this is way less work. You would have to regularly check and change your oil and if it malfunctions you get oil all over your hash. But the oilless version also costs an extra 2000-3000€ depending on the manufacturer.

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This brings us to our next point: how do I use my freeze dryer? We already told you about the pump maintenance, but this is not the only thing you have to check before using your dryer. Before each cycle you should check the drain valves. Let out all the drainwater and the close it again tightly, because if you forget that you won't be able to produce a full vacuum. Now we clean the inside of the dryer and remove any leftover ice (if you haven't defrosted). We connect the pump to the dryer and inspect the doors if they fit snuggly on the sealing part that guarantees the ability to pull a vacuum. If we prepared our trays right and they look like this we can load them into our FD and start the process. This will take about 24-36h depending on how thick your hash layer is

To check if it's dry, take a card and move the hash around a bit. If it behaves like sand and has no clumps left it is dry and ready for smoking or further processing

Drying Hash without a freeze dryer

This method is mostly used for home made hash for it's cheaper tools. As said in the washing guide you should immediately freeze your hash after collecting until it's a hard brick. This gives you the opportunity to separate the individual trichomes better in order to get more surface area for water to evaporate. You need a temperature controlled room with about 30-40% humidity at all times! The room should also be clean to insure no mold spores get into your hash and multiply.

Now you take baking paper sheets and lay out your drying area. You can use baking trays in an open room setup or put it in a clean (!) pizza Box as the cardboard will pull out even more moisture. Then take your frozen hash bricks and grate them either with a microplane or a siever finely over the paper. Be careful to really spread it out good as clumps may lead to inconsistent drying. Wait approximately 1-2 weeks and check regularly for dryness.

How do I calculate my yield from flower to Hash

Now that we weighted out dry hash and have a g number we can simply take the % Value e.g. 5g Hash out of 100gs Flower would equal a yield of 5% These values differ greatly between WPFF and cured Hash as you have to subtract the water weight from the fresh frozen flower. The fresh flowers are about 75-80% water so 5% is a high yield compared to cured hash yield where 20-25% would be a good yield. Good fresh frozen yields are between 3-8% depending on the strain, 8% being the top of the top. Most times these numbers are only achieved with selected cuts asa lot of strains yield <1% For cured Hash a good range is between 15-30% return, but this also requieres a hash specific cut

More Hash-Education

If you want to learn more about Hashtech and Hash culture visit my buddy over at TheHashishInn and listen to his Podcast

What does colour say about my Water-Hash

This is a very controversial discussion in the water-hash space and I hope to not step on anyones to toes, but this needs to be said. Colour doesn't affect the quality of Hash! Most use this phrase but forget to say that, while this is true, there can be bad dark hash. Sometimes it's not a sun ripened or cured resin, but just contaminants. This is a important factor to consider when buying Hash. You have to look really close and do the ultimate test: how does it smoke! No real hash connoisseur cares about the colour when the flavor, smoothness and high is right and so should you. Some people go as far as harvesting way to early in order to get the whitest hash, while it only leads to a speedy, weak high and nearly no flavor

"Dark" or better said amber hash can come from strong UV-Radiation, CBN-Content or long air exposure which oxidizes the compounds in hash

How is Hash rated?

Most of you will have heard about 6 star hash or lower quality, but exactly is that measured? Tbh the star-rating system is used quite liberally so a lot of "six-star" hash is worse or not even close to being fullmelt. To give you a quick overview of how it should be used and a measurable alternative shall be given here

The star rating system

1-2 Star

This grade is mostly used for edibles/RSO or further cleaning processes as it's from low quality. You can identify it by it's green/dark colour that stems from the high content of contaminants. You will typically find this in the 180u or 25u bags

3-4 Star

We use this grade of water-hash for edibles on the worse end and on the better end of the quality spectrum we press it to rosin as this will still yield a proper, terpy oleo-resin. Typically found in the 150u Bags and characterized by it's light colour but missing meltyness

5-6 Star

This premium product will be processed to high-end-rosin on the lower quality spectrum and the best of the best will be left as it is if you have a market for melt. If not, you can also press it and sell it for the maximum rosin price, but beware I will call it a sacrilege to press such high quality melt haha True fullmelt shouldn't leave residue in your banger. A tiny bit is normal as the trichome heads still have their waxy cuticle and this "burns"

The gravimetric measurement, a good alternativ?

As you might have noticed the star-rating leaves a lot of room for interpretation and therefore is unprecise. Fletcher from Archive Genetics wanted to adress this issue and bring in a measurable test. As he was the one to bring freeze dryers into the hash industry, he has a lot of credibility in this regard.

His method is to use an enail, take a 15micron metal screen and weight it before. Now you put exactly 0,1g of your Hash on the screen and put all that on the enail until no oil is left. Then take the screen and weigh it again. Now subtract the previously taken screen weight and there you have your residueweight. This also can be converted into a %melt value which should be used to categorize the Hash Visit his Instagram for fire flower and melts

 

 

 

 

 

 

Sieh dir diesen Beitrag auf Instagram an

 

 

 

 

 

 

 

 

 

 

Let’s end the star system for water hash, solvent less, ice wax, and move onto a system that accurately grades how “MELTY” your hash really is. just simple gravimetric analysis and it’s relatively accurate for the end consumer’s needs. Tag your favorite hash maker in this post! @trichomeheavyextracts1 @k3nnnwall @cannabisreverend @kush4breakfast @710labs @3rd_gen_familyfarm @resin_ranchers.pdx @hashcamp @icedoutextracts @beezlegardens @theccc420 @dannyflavors @shane_omac831 @pureoptions @jungleboys @jungleboysfullmelts @humboldtorganiccollective @michigan_made_melts @icextract_equipment2.0 @cubangrower @tharealogkushman @eldaggy @turtle_trees @megaraw_melts @voodoomelts @solventlessismore @puaextractions @fullflavaextracts @golden_trichome_extracts @dankczar_icewax @connected.california @alienlabs @returnofthealien @ogkushbreath @therealogkushstory

Ein Beitrag geteilt von Archive Seed Bank (@archiveseedbank) am Apr 19, 2020 um 6:09 PDT

What can I do with my finished product?

This depends on your needs, but most people dab/press it or use it for high end edibles. If you process it to rosin you can also make carts with it or fill preroll "donuts" with it as this is quite popular right now The leftover washed flower can be freeze dried and be blasted/alc washed to get every cent out of it. If you already separated all trichs you can use it as mulch for your garden aswell. I hope you learned something new and had fun doing it. If you have any tips/tricks or corrections you want to be included, feel free to leave a comment or shoot me an email as I want to improve the quality of my hash making aswell and you never stop learning Happy dabbing everyone PS: Sorry for my english, I'm not a native speaker :D

Disclaimer: This tutorial is intended for legal use in legal states only. We do not condone any illegal activity

Sources

1. Dr. Ethan B Russo; "Taming THC: potential cannabis synergy and phytocannabinoid-terpenoid entourage effects"; (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3165946/)
2. Analytical 360; „Bubble Gum Hash Rosin“ 16.06.2016 http://archive.analytical360.com/m/concentrates/565901I
3. Iceextract; ICExtract Bag Singles and Kits 5Gal V3 https://icextract.com/icextract-bag-singles-and-kits-5gal-v3/
4. Pollonator/Mila Jansen; MEDIUM ICE-O-LATOR® 7 BAG SET; https://pollinator.nl/product/medium-ice-o-lator-7-bag-set/
5. GoPurePressure; Bubble Now 220 Micron Work Bag https://gopurepressure.com/collections/bubble-bags-machines-bubblehash/products/bubble-hash-bubble-now-220-micron-work-bag
6. homeairguides.com; How To Choose An Air Conditioner (Step-By-Step Guide On Picking The Right AC Unit; https://homeairguides.com/air-cooling/how-to-choose-an-air-conditioner-guide-on-picking-the-right-ac-unit/
7. Deborah Hucht; Kühlleistung berechnen: Welche Klimaanlage eignet sich für welche Raumgröße?; Kühlleistung berechnen: Welche Klimaanlage eignet sich für welche Raumgröße?
8. Ab Hanna; "Hash Rosin 101: Lessons from Experienced Solventless Extractors"; https://hightimes.com/guides/hash-rosin-101-lessons-from-experienced-solventless-extractors/
9. PurePressure; Andrew Ward; "How Bubble Hash is Rated (1* to 6*)"; https://gopurepressure.com/blogs/rosin-education/how-bubble-hash-is-rated-1-to-6
10. Titelbild: CubanGrower/https://www.instagram.com/cubangrower/ / https://www.instagram.com/_thevillage/
11. "HOW DO REVERSE OSMOSIS SYSTEMS WORK?"; (https://kraaiwelldrilling.com/how-do-reverse-osmosis-systems-work/)
12. "Different Types of Terpenes in Cannabis"; (https://tetrahorizon.com/2020/07/10/different-types-of-terpenes-in-cannabis/), TheProfessor

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by Max

Plant Tissue Culture - the future of Cannabis clone production

featured image by the one and only @kandidkush

1. Introduction

In recent years, automation and standardization of the crop production process are becoming increasingly important. Especially in the production of cannabis, it is crucial to have uniform and disease-free planting material in order to reduce any plant protection measures to a minimum and to ensure a final product with consistent quality. For this reason, more and more growers decide to source their seedlings from in-vitro propagated or tissue cultured material. But what does In-Vitro production and tissue culture even mean? And how is it done with cannabis? That is what we want to explain in this Article.

In Vitro is Latin and means “in glass” or “in the glass” therefor when according to plant production, it is a technique that uses glass containers such as petri dishes and test tubes as a controlled artificial environment for the propagation of plantlets. This contrasts with in-vivo ("within the living") and in-situ ("on site") production, which are commonly used in horticulture.

Within in-vitro production there are various methods and ways to propagate the desired type of explant. Tissue culture and micropropagation are two terms that you will most likely come across while looking closer into that topic. But what exactly is the difference between those?

The main difference between micropropagation and tissue culture is that micropropagation is the production of a large number of plants from a small amount of plant material, whereas tissue culture is the first step of micropropagation, where plant cells are grown in an artificial medium to develop them into a large number of plantlets. In addition, micropropagation requires tissue culture for the propagation of plantlets.

As the global market advances to a greater reliance on plants for active ingredients, delivery of consistent and pathogen free plant material is crucial. Anyways, in the production of medicinal or recreational cannabis, the use of these techniques is not yet widespread, as cuttings can also be produced via "normal" vegetative propagation but, the higher the degree of automation and the producers' demands for cleanliness become, the more this method comes into focus. In addition to the possibility of producing clean plants, in-vitro culture is also suitable for space-efficient growing, improved yields due to vigorous plants and for saving production costs.

However, the in vitro cultivation of plants is not only suitable for propagation but is also of great interest for research and especially for breeding. For example, pathogens can be eliminated from plants, mutations can be created and sterile backups of rare or hard-to-retain plants can be secured. There are many ways to utilize tissue culture and other biotechnological methods and there are even more to be discovered.

2. Basic Terms for In-Vitro production

2.1 Medium

2.1.1. What's Agar Agar?

Some of you might know it as a plant derived gelatin substitute and that's nearly the same purpose we will use it for. The thickener is based on a galactose containing extract taken from red-purple marine algae. This brings the typical nutrient solution to the right consistency that supports vigorous growth, but gets easily penetrated by the roots to insure the best development possible. In order to generate the perfect Agar mixture we have to add it in a ratio (depending on the source) 1-2:100. This leads to a nearly complete solid medium. If you want to cultivate in anaerobe/liquid conditions, you can simply half the concentration.

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But why not just use gelatin instead?

That's a good question, but there's a simple answer to it. It's edible to bacteria and most other microbes so you would have to refill every plate after a short time. The nutrient dosing and validity of the experiment undertaken would be in danger as you would have to add the gelatin to the feeding chart of the microbes that are cultured.

It comes in different forms, but the most common are flakes, that need to be mixed with water und constant heat application until it's fully homogenized. These flakes are easy to store and quite cheap to get. If you have the necessary budget, there are pre-mixed/pre-sterilised versions, mostly bottled up, that can be used instantly without waiting for the hot agar mix to cool down. The cooling process can take up to 30min per batch before use.
Not only the distribution form differs but also the additives. Many agar mixes have added ingredients for certain purposes as this is used for all types of cell cultures from human cells to the infectious disease causing Escherichia coli. 

The potato-dextrose agar for example is the most used form for fungi especially for varieties like Botrytis cinerea also known as the common grey mold. Another one is blood agar, which as the name already says, contains animal blood in order to study special microbes. For tissue culture the most common type is the standard galactose agar in combination with a nutrient matrix and a hormone, depending on the phase.

The standard nutrient matrix in nearly all cell culture work is the Murashige-Skoog-Medium, which contains simple amino acids, nutrient salts and photosynthesis products (saccharose) as the plant can't photosynthesis in most cases. Inositol is also added in order to mitigate plant stress, strengthen cell walls and phosphate storage.

2.2 Hormones for plant tissue culture - is that even safe?

Many myths rank around plant hormones, which are mostly "feared" by the Cannabis community as it only has found real application in large scale agriculture. They get mixed up and thrown in the same bucket as GMO and Glyphosate. Not that those would be bad substances but the social stigma around them is strong.
The hormones on the other side are nearly identically to their natural/plant counterparts, which control nearly all the physiological responses of the plant to outside influences like the movement of the sun or the alignment of the plant to the gravity

2.2.1 Auxin

The first of these phytohormones (hormones in plants) is responsible for a multitude of responses to environmental stresses. Most auxin derivates in the plant and made by humans are based on the Indol-3- acetic acid. This compound is mostly synthesized in the apical region refering to the main growingtip of the plant. From there on it gets transported via the phloem and cell-to-cell by the PIN 1-9 transporterprotein.

The basic effects include wound response, apical dominance (the main branch is the tallest one), photo-/gravitropisms and it plays a important role in fruit development. But lets dive a bit deeper in the interaction of Auxin and the use cases of it.
The first practical application would be induction and promotion of root growth. This is the most used purpose and nearly all commercial rooting gels contain it. But not only in Cannabis, but also the floral industry uses it predominantly in their rooting SOP.
We also use this by giving our explants (plants in in-vitro conditions) a certain amount of it in order to achieve the highest rooting rate possible.

But as with everything in life, there is a limit to whats good for the plant. Excessive application of this substance can result in growth inhibition and plant death. Weedkillers like 2-4-D are based on the synthetic counterparts of auxin like indole-3-butyric acid. But not only damaging substances also "good" ones like common rooting gels are based on synthetic auxin.
"Natural" Auxin can be found in variety of plants shoots especially the ones of the willow tree.

2.2.2 Cytokinin

The next hormone in our repertoire is the counterpart to the previously mentioned Auxin. This compound focusses on the lateral growth of both roots and shoots. To be more exact the ratio of Cytokinin:Auxin is the driving factor in the morphology of the plant. They not only contradict each other, but also work together when influencing cells. When only Auxin is applied cells elongate and become big, but won't expand or differentiate. Same takes place when the ratio is 1:1. This is used for Callus multiplication or expansion.
Complementary to Auxin it is synthesized in the roots and travels to the shoots via Symplast and Apoplast. Production of this chemical is regulated by two types of response regulators. One being the B-Type and the other being the A-Type. Both are regarded as transcription factors as the either activate or stop the production of Cytokinin via influencing the transcription of the corresponding gene loci.

Using this knowledge we can manage the morphology of the plant into growing more squat thus increasing internodal stacking and use of space.
But not only the space can be used more efficiently also the time saved by breaking seed dormancy can save time when germinating seeds for a phenohunt. The dormancy is induced by abscisic acid and by increasing metabolic activity the cytokinin decreases the level of abscisic acid in the seed. It's commonly used in plants that have a hard time germinating.
The next effect might be especially interesting for the photographers and florist under you. Cytokinins can delay senescence in other words the decay of the plants. This is achieved by increasing synthesis and slowing decay of certain proteins. Also nutrients are drawn into the treated area from nearby tissue. It's suspected that an enzyme is responsible for these actions, but no scientific consensus has been reached yet.
When using Cytokinins you have to differentiate between the plants own, adenine based, Cytokinins like Kinetin and Zeatin. Others have been found outside of plants, but they are based on Phenylurea. These exceed the effectivness of the Cytokinin in certain plants. Examples for this variety are TDZ and Diphenylurea, which are widely applied in agriculture

2.2.3 Gibberellic Acid

One of the most interesting phytohormones is the counterpart to abscisic acid. Exogenous GA3 (short version) is also responsible for breaking dormancy where it activates indigenous synthesis of more GA3, which adds to dormancy breaking factors. These are hypothesised to be a combination of growth promoting hormones (GA3, Cytokinin, Auxin etc) and reduced nutrient storage in the endosperm. Enzymes, primarily α-amylase, lead to the processing of sugars and other storage units. Another restricting factor is the Cuticula strength of the seed.
The application on an adult plant leads to a big stretch in all shoots. This is principle is also used in plant tissue culture and for virus cleansing. But this has to be done with the utmost focus on the concentration of GA3 as a high amount leads to different sex expression. This mechanic is commonly used in feminized seed breeding as it "reverses" a female plant into producing pollen with female Genome. In even bigger concentration it can lead to the complete sterilization of the specimen. Most fruit growers like in the citrus industry use the mechanic to suppress undesirable seed development

That said, when a trained horticulturist with a strict, thought through regime applies GA3 in flower, it can increase inflorescence mass and trichome development. For homegrowers and people that don't have access to these chemicals, Kelp/Seaweedextract is a perfect allrounder as it contains a variety of growth hormones aswell as Macro-/Micronutrients.

The compound was discovered by japanese scientist while researching the Gibberella fujikuroi fungi. Cytokinin is a secondary metabolite to the pathogenic pathway of the fungi on rice plants.

2.3 Basic Tools for Tissue Culture Propagation

2.3.1 Laminar Flow Hood

It's one of the most important, if not the most important part of a Culturelab. The Laminar Flow Hood (LFH) insures that all steps are done in a steril environment. There is a differentiation between normal Flow hoods and special models for working with microbiological organisms. The first doesn't filter the incoming and outgoing air, leaving the worker exposed to aerosols and microorganisms.

That's why we use the second model in most cases. This LFH got a specialized HEPA-Filter before the air enters the Hood. After getting in it can be distributed in two ways. Vertical flow hoods and horizontal flow hoods can be differentiated easily as word explains itself

All work including the plant have to be done inside to keep out potential contaminants. It's therefore of vital importance to wipe down all areas inside with ethanol (70%) and clean the outside of culture vessels aswell when introduced in the steril environment. Gloves and hair-protectants are a must, if mass-propagation is planed. At home you can easily get away with disinfecting your hands and arms up to the elbow. Another concern is your breath as it often contains bread-mold spores and a variety of microorganisms that thrive on tissue culture medium.
If a infection takes place and you have no copy/other option, you can add antibiotics to the medium, but be very careful. Most higher plants are more resistant to these compounds than the microorganisms, but the optimal balance is very hard to reach. Another factor is the difference between every plant genome/phenotype thus a cultivar-specific test would be necessary before adding antibiotics.

2.3.2 Autoclave

Speaking of sterilisation, we will subsequently be lead to autoclavs. These laboratory certified pressure cookers are an essential tool for cleaning glassware and used culture vessels. They come in different sizes and forms, which you can choose specific for your application.
Despite the different shapes, the mechanism is the same. The chamber is evacuated via a Vacuum pump in order to simplify the sterilisation progress. It gets filled with hot steam and thus put under a high pressure to increase the cleaning efficiency. After 15-20min the optimal cleansing effect is achieved, which means that the content can be removed. Be careful not to touch it with your bare hands as it's still really hot.
You should also size the autoclave big enough for the operation, but not too big for your batchsizes as this will lead to unnecessary cost for energy.

2.3.3 Magnetic Stirrer

This device is rather simple and doesn't need much explanation, but it's still vitally important that we talk about it. You probably wonder why and we will will explain it shortly. Not the stirrer per se needs explanation, but rather how to use it and what for. It's the base of a good tissue culture lab as it's used for mixing and homogenizing of the medium components that we discussed earlier.
Each component has a specific boiling/inactivation point that you have to consider when mixing.
The goal is to find the right compromise between solubilityspeed and retention of effectiveness. Some compounds like hormones derivates have to be added after the mixing + autoclaving of the medium as they can't even withstand the autoclaving procedure.
For these substances you have to consider other ways of sterilising them such as syringe filters or specific solvents.

It enables the production of large amounts of media as it's fully adjustable in rotationspeed and temperature, if you have a heated version. Be careful to pick a appropriable sized stir bar as this determines the homogeny of your mixed substance.  But even with a large stir bar the stirrer has it's limitations when the mixing materials are too viscous. If that's the case, we would advise a mechanical alternative as they can plow through it better. Keeping that in mind, you have to pour your hot, mixed medium out of the vessel before it cools down as it gets really hard to remove the bar after.

2.3.3 Consumables

This chapter is for items that are important, but didn't get a separate chapter as it would go beyond the scope of this article.
Firstly we have your PPE like gloves, masks, Labcoats and hair nets. These are especially important, if you have a large team and thus many vectors of contaminants/pests. But keep in mind that PPE doesn't eliminate the necessity of disinfecting the whole work environment before and after you do explants in it.
Another vital point are the culture vessels. These are the new homes for your little clones and need to have a large enough volume to accommodate them. This refers especially to the amount of medium that a plant needs in its stage of growth. There are mayor differences from cultivar to cultivar so the best way to go about it, is to  experiment with it and keep tight journals in order to create SOPs. This takes a lot of time, energy and labour so be mindful of that, when trying to establish a plant tissue culture production.

Our last important item is the scalpel.  You can find many different forms of it for specific applications, but we recommend that you go for a more environmentally friendly version by using a scalpel holder with exchangeable blade. Using this will decrease your waste of material and money as the amount of cut plant material in an TC production puts a great stress on the blade thus making it dull. The alternatives would be to use whole single use scalpels, but the produce large quantities of plastic that we don't want or high end versions. The later stays sharp for longer, but not long enough to make up for the huge price difference.

2.4 Standard terms for plant meristem tissue culture

In order to understand the basic mechanics of saving genetic material and growing callus cultures we have to take a look at the structure of a growing plant tip. This is the location we're using to explant a meristem from. But what is a meristem?

The term comes from the ancient greek word merisein, which means divide. It was used by the swiss scientist Carl Wilhelm von Nägeli to describe the undifferentiated, multiplying Cells that made up the growing tips of the plant. The apical meristem is the highest growing tip in the plant and sits on top. The counterpart of this region is the basal meristem, which sits of the tip of the roots as you might have guessed.

The plant builds new meristemcells in the top middle parts as the lower parts differentiate into the predetermined functional parts. Some go to be Parchenym cells, some are going to be part of the vascular system of the plant. Knowing this, we can extract these cells before they're differentiated in order to get totipotent cells (cells that can be any part of the plant). These are full of potential and are kept in a cryofreezer to be saved for later usage, as backup or as basis for mass cell production in a bioreactor.

3. Basic mechanics of mass micropropagation

To get an idea of how micropropagation could benefit your growing operation we're gonna draw a little plan of how it could look like.
First of all we need a small motherstock from which we can take our microcuttings. These are about 2,5cm/1 inch long and at best, taken from a meristematic region.
Then we plant it in a multiplication medium under sterile conditions in order to avoid contamination and the subsequent demise of our plant matter. After 2-3 weeks we should see 2-4 new shoots coming from our tissue, which we can also cut up into separate tissue samples. The we put them in individual containers with the same medium mix again and repeat the previously mentioned cycle. Even thought there will be loses due to contamination, the replication-rate is extremely high due to each new tissue replicates itself 2-4 times. If we calculate this through, we can see that with minimal space use, an enormous plantcount can be achieved in a short period of time.
After we finished the replication cycle, we can move onto the rooting phase. This uses a special medium containing auxin derivates, which induce the production of roots on our previously made shoots. After 2-4 weeks again, we have to inspect the plants accordingly and if they're sufficiently developed, we beginn the hardening phase.
For this part of the productioncycle the plants get translated in the medium of choice that will be used later on in production. Most growers will use rockwool as it has excellent water holding capacities, but also coco or small soil pots can be used. Light plays a huge role now and has to be increased in order to acclimate the plants to the later vegetative conditions.
The humidity also has to go from the comfy 80-90% RH in the boxes down to the values of your vegroom.
This was a quick overview of the process and as you could see, there are many obstacles along the way, but in the end it's not that hard to achieve at home. For this exact purpose we will compare DIY kits that can be bought online with a home made microlab.

4. What are ready-made home kits?

There are some suppliers on the internet who offer hobby kits for micropropagation. Even if these kits are suitable for experimenting and for first experiences, they cannot replace a professional laboratory, because similar to mushroom cultivation, a sterile working method and environment must be guaranteed to avoid failures. In addition to all the materials and equipment, professional application requires trained personnel and constant quality control of the work process.

5. Future vision

As cannabis production becomes more and more automated and the costs of labor, electricity and rent continue to rise, we at research gardens see in-vitro production as a key advantage for companies that want to remain sustainable in a rapidly evolving industry. Due to the prospect of a long-term reduction in production costs as well as a simultaneous increase in product quality your company will get an advantage against your competitors. Particularly in pharmaceutical operations, a consistent quality of the plants and ingredients is indispensable. For the recreational market, however, this is just as important since similar requirements exist here as well.

Interested? Then let us start working out a concept for an in-vitro laboratory in your company today! Send us an e-mail to info@research-gardens.com

Sources:

 

1. "Communication by Plant Growth Regulators in Roots and Shoots of Horticultural Crops" by Anish Mallad; Jacqueline K. Burns; August 2007; HortScience: a publication of the American Society for Horticultural Science 42(5) (https://www.researchgate.net/publication/237379591_Communication_by_Plant_Growth_Regulators_in_Roots_and_Shoots_of_Horticultural_Crops/figures?lo=1)
2. "Hot Plate with Magnetic Stirring: 6.7"x6.7" SS Plate Max.300C - SH3"; MTI Cooperation; (https://www.mtixtl.com/hotplateswithmagnetcstirring67x67stainlesssteelsurfaceupto300c-eq-sh-3.aspx)
3. "Sterilization cycle phases for a steam sterilizer"; rsd-engineering.com; (https://www.rsd-engineering.com/steam-sterilization-cycles)
4. "Laminar flow hood/cabinet- definition, parts, principle, types, uses"; October 1, 2020 by Anupama Sapkota; (https://microbenotes.com/laminar-flow-hood/)
5. "Baum HD"; Dreamliner (https://wall.alphacoders.com/big.php?i=612687&lang=German)
6. "CULTURE MEDIA AND ITS TYPES USED UN MICROBIOLOGY LAB."; milan; (http://biologyofmicroorganisms.blogspot.com/2010/09/culture-media-and-its-types-used-un.html)

7. "[Tuesday Scoop] Puzzlement Between Agar-Agar And Gelatin"; Namrataa Mahalley; (https://bulbandkey.com/blog/tuesday-scoop/puzzlement-between-agar-agar-and-gelatin/)

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by Max

The EC value: importance for (hydroponic) cannabis cultivation

The EC value is a physical unit [1] that indicates the electric conductivity of a substance.
Knowing the EC value, we as growers get an overview over how many nutrient salts are dissolved in a nutrition solution.
This is possible because nutrient salt molecules break down into their individual ions in a watery solution and generate electric conductivity in it.

These ions have free charge carriers. More free charge carriers in the sense of unoccupied electron sites or excess electrons far from a charge equilibrium, ensure a higher electrical conductivity and thus, a higher EC value of a solution. The chemical background however is not that important for us gardeners, because there are easy to use EC-measuring instruments for practical purposes. Later in this article we will dig deeper into the scientific backgrounds of the EC value, but now it's time for some hands on information.

Measuring with a EC device you will be most probably confronted with microsiemens per centimeter (1 ms/cm) or sometimes also PPM (parts per million) or tds (total dissolved solids). It has to be said, that the expression of the electric conductivity in ms/cm is the most accurate for horticulture applications. You'll find usage of this unit mainly in Europe, while tds and PPM is very common in the US.

The most important use of measuring the EC value is during the mixing process of your fertilizer stock solution, that irrigates your plants. This ensures that there's always a proper basis for your plants nutrition needs.

The second important application of measuring the EC value is related to monitoring the health of your growing medium. With measurements of soil EC, Rockwool EC, Coco EC or recirculating nutrition solution EC you can check the state of health of your medium. This is crucial for a successful grow, as the amount of dissolved fertilizer salts in your medium affects the oxygen concentration in your medium and also can lead to nutrition burn, which slows down or stops plants growth. Especially in the flowering stage high oxygen amounts in your medium are very important for a vigorous grow of fat flowers.

The following EC values are a good starting point for EC values during the cannabis life cycle (based on personal and common research experiences):

Medium Crop Steering with measuring input EC and drain EC

The EC value gives you a great starting point for the mixing process of your stock solution / fertilizer mix. Whenever you irrigate your plants, especially in hydroponics, a well mixed fertilizer solution fitting to the needs of your current plant development stage gives a great basis for a successful grow.

But it gets more difficult when it comes to the EC value of your medium. This is because depending on a plants individual nutrition uptake behavior the EC value in the medium can rise over time, even if you always give the same (low concentrated) EC fertilizer mix every time. This is the reason why I would recommend also measuring the medium EC value. With this technique you're able to "steer your crop" nutrition-wise.

How is crop steering be done? Most basic rules for crop steering with measuring the EC value of your medium.

If you irrigate your plants (automatically or by hand) you should aim for a little runoff / drain after some irrigation events. This excess drain water you can collect in a small beaker glass and then measure it with your favorite EC measuring device. If you measure higher EC values in the drain than in your nutrient solution, you can conclude there have been happened a salt built up in your medium. This indication gives you the sign to lower the fertilizer concentration in your stock nutrition solution for the next time(s) to slowly lower the medium EC value over the time.

On the other hand, if you measure your drain and you realize it's EC value is lower than what you apply with your stock nutrition solution, it gives you the sign that your medium and plants need to be irrigated with higher fertilizer salt concentrations.

If you never measure your drain, you also will not recognize any salt built ups beforehand and could be surprised by deficiency or nutrition burn symptoms when it's already too late. Especially when growing organically in soil you're not be able to react fast to rebalance your soils conditions and it can be already too late for saving your crops potential yield. [2]

Be careful: The EC value alone tells you hardly anything about the abundance of specific nutrients in your fertilizer solution or soil

And it even gets more difficult: Depending on a plants nutrition uptake behavior, a medium can also develop nutrition imbalances. Because as plants require 17 different chemical elements for healthy growth, plant nutrition is a bit more complex than just measuring the EC value. While you can be sure that fertilizer component ratios are in a righteous ratio when mixing A + B components following the fertilizer scheme with your plain water, it's a more difficult thing when it comes to these ratios in the medium or soil.

Without a proper lab analysis of your medium you will not know, which salt is missing or if there's too much of one in your solution. Luckily this can be roughly analyzed when combining medium EC measurements with medium pH measurements. In short: Different nutrients have different pH levels. Ammonia - a plant available form of nitrogen - has a very alkaline pH of 11. Ammonium dihydrogen phosphate (ADP), a plant available form of phosphorus, has a pH of 4.2 in a 5% concentration. So for example when the pH value of the nutrition solution when irrigated is 6.0 and the drain pH is 5.5 we could assume that plants took up more alkaline nutrients like nitrogen and less of phosphorus as the pH of the medium got lower. This advices us to give more nitrogen and less phosphorus next time when irrigating. But that should be it for now regarding pH. More in this in one of our upcoming articles.

Not all dissolved salts in a watery solution help plants grow

Since the EC value does not only measure the required nutrient salts for plant growth, it shows us only a vague picture of what is going on in any solution nutrient wise. Part of the measured EC value are also other salts that are not usable by the plant such as sodium chloride, which could be more described as toxic or destructive for plants. This is the reason why professional gardeners work with laboratory soil or medium analyses in order to get to know which single types of salts make up the total dissolved salts in a solution or medium. With this knowledge in mind, one can see which nutrients are missing, which need to be added or which salts are too much in a mix. In the high end professional field, this can be automatically controlled by a fertilizer computer and a closed nutrient solution circuit. In most cases, these measurements are not integrated in irrigation computers and are done from time to time in the lab with help of a so called photo meter.

As this article focuses on the EC value and not plant nutrition in all aspects, we will proceed with this topic some lines later.

What is important to keep in mind at this stage is just the fact that whenever you measure the same input and output EC does not automatically mean, everything is fine with your medium bound salt concentrations. Because it can be still be, that there are some hazardous peaks for one or another specific salts.

The importance of tap water EC value

The previous paragraphs are most important to keep in mind when operating with tap water. Tap water can already have EC values up to 0.9 ms/cm - mind you, for the most part these dissolved particles are not useful but toxic nutrient salts which nevertheless strongly increase the EC value of your nutrient solution or your substrate over time (to your plants disadvantage). As the upper table shows, younger plants need lower EC values under 1 ms/cm. So 0.9 ms/cm tap water full of toxic salts like sodium chloride provide no room for nutritional salts from your fertilizer bottles and could lead to sustainable damages at your plants.

A helpful solution for this very problem comes in handy in form of reverse osmosis filter units. There is a paragraph at the end of this article about such devices. Whenever you start a grow at a new destination, I highly recommend to measure the tap water there. If the tap water EC is over 0.4 ms/cm it's best for your grows success to invest in a reverse osmosis unit.

Plant physiology background on EC value:

Nutrient and water uptake via the roots depends largely on the EC gradient between the substrate / nutrient solution and the plants internal salts concentration. The goal is to achieve an equilibrium of nutrient concentrations between root cells and the nutrient concentration in the medium. To give an example, an equilibrium would be achieved, if both solutions in the root cells and substrate each consisted of 99% water and 1% nutrient salts. Or 98% / 2%. Important is just, that it's close to each other. If the ion concentrations of two solutions separated by a semipermeable, sieve-like membrane differ (e.g. 2% to 5%), the aim would be to equalize the concentrations, which is not to be confused with a pure mass or volume equalization based on simple pressure differences. That would be "just" diffusion.

Osmosis

The most important type of mass transfer for the EC value is called osmosis. In this process the individual components of a solution, in our case the water on the one hand and the nutrient salts on the other, do not move proportionally from one cell to the other, but always strive for a homogeneously concentrated solution in neighboring cells.

Imagine cell walls as a semipermeable membrane like a sieve, that lets small water droplets pass. As we know that two neighboring cells strive for same salt concentrations, there are two ways to achieve this: The cell with the lower salt concentration has to let some water flow to the neighboring cell to get both cells salt concentrations to the same level. Or the cell with the higher salt concentration sends some water to the cell with the lower concentration. And all this happens automatically, as salty water has no hurdle to flow through the semipermeable cell membrane. FYI: The semi-permeable membrane is semi-permeable, because it lets salt enriched water freely pass, but not bigger molecules like glucose. Transporting these bigger and more complex molecules has to be done actively with the help of some plant energy in the form of ATP. Water molecules in comparison travel between cells passively without the need of extra provided energy just according to different salt concentrations. The motor for water transport in the end is the transpiration suction of the leaves.

Effects of high medium  EC values with low plant EC values

If the nutrient solution or medium has a higher salt concentration and the plant in its root cells has a lower salt concentration then the root cells lose water to the nutrient solution as a result of this condition with simultaneous nutrient salt uptake and the cells can dry out. This occurs because a balance of salt concentrations in the cell sap and the surrounding nutrient solution is sought, with water leaving the root cells for dilution of the saltier medium or nutrient solution. At the same time, nutrient salts are drawn into the plant cells.
This causes an excess of nutrients in the plants cells and at the same time a shortage of water in the cell. Plants can die from this.

Effects of low EC value of the nutrient solution or medium with high EC values of the plants

If the salt concentration in the root cells is somewhat higher than in the nutrient solution, everything is fine. Because more salts are stored in the root cells, there is a gradiation balance in favor of the water in the direction of the root. This means that in favor of achieving a balance the saltier root cells are accordingly diluted with water from the nutrient solution in order to establish the concentration balance.

All in all, lower EC values are better for healthy plants. Because it's way easier to increase salt levels in plant tissues than get rid of too much salts in a plant. But if the EC value in the medium is too low for too long, that also could be a problem.
For example, if the EC value of the root is very high and the EC value of the nutrient solution is super low, too much water may be absorbed by the plant, while the uptake of nutrient salts doesn't happen. This then manifests itself in pale leaves, less dense and large flowers or overly slender growth. A classic case of under-fertilization.

In the plant, a high EC value of a nutrient solution manifests itself in wilting and hard leaves, stunted growth or even cessation of growth. This is a classic case of over-fertilization.

Role of plant EC value for photosynthesis and cell respiration

The challenge of the matter is the successive increase of the EC value in the plant - which also allows us to increase the EC value of the medium gradually without upsetting the osmotic equilibrium of the plant. [4]

The EC value in the plant increases over time because it draws water vertically through the xylem (water pipes) of the plant. The water carries the nutrient salts upwards, evaporates due to heat and leaves the salts back in the plant cells because they cannot leave the plant like water vapor can do. This is either due to their molecular size, and/or relative density/weight in comparison to water vapor. Thus, the nutrient salts keep stored in cells and get transported to where they are needed. In addition to the Xylem water pipes, there are other vascular bundles in the plant, called phloem, which can only transport nutrient salts and sugar throughout the plant. [5]

Basically, the plant first transports water and dissolved nutrients to the photosynthetic organs, mainly leaves, to perform photosynthesis. The main product of photosynthesis, the energy-rich glucose, is then transported via the phloem from the leaves back to the roots in order to be able to perform the second important metabolic process of plants: cell respiration.

The glucose production by photosynthesis is one of the plants first important metabolic processes in the energy supply chain of a plant and is made out of light energy, CO2 and H2O.

Glucose then gets transported down to the roots to be saved down there and gets destructed again to carbon dioxide (CO2) and water (H2O) in the process of cell respiration. During this process of cellular respiration the plant get a lot of chemical bound energy namely in the form of "ATP". FYI: Cell respiration plays a mayor role in production of fat buds. Because during the built up of flowers a lot of higher molecules like glucose has to be transported through the plant - for this there is big need of a lot of ATP, which is required by active plant transport mechanisms. In comparison to nutrition salts, glucose cannot pass semipermeable membranes.

The bottom line is, that cellular respiration works with the results of photosynthesis and vice versa.
Difficulty: Both processes take place at the most distant organs of the plant (leaves and roots). [6] I would like to show you a small sketch for clarification:

The plant increases its own EC value slowly but continuously. Time is the determining factor.

The plant in the vegetative stage will grow very much in height and thus creates many cells and with some weeks of time, the plant can still grow in height and width and also lignify, continuously making new space for future nutrient deposits.
Thus, the EC value, the percentage of nutrient salts in the plant at the beginning of growth, does not increase significantly, more does the number of cells and the absolute amount of nutrient salts increase.

Later in the flowering stage, when the plant starts to grow in thickness and width, there aren't created so many new "superstructures" like before, but the existing cells get pumped up with nutrients and more important higher molecules like glucose. At that stage, of course, the plant can use a lot of nutrient salts, because the plant no longer concentrates it's energy on height growth and cell division, but on the pure accumulation of flowering mass and cells to increase it's own reproductive probabilities. This all happens in a fairly straightforward manner.

A plant grows, when the photosynthetic activity is higher than its cellular respiration

Cellular respiration is done by roots in the dark with the help of oxygen. A plant grows, when it undergoes cell division [3] and the individual organs of the plant, such as leaves and shoots develop.

It's essential to form enough photosynthetic organs (leaves), which on one hand provide the necessary transpiration suction to take up water through the roots, on the other hand mainly for the conversion of sunlight and water into glucose, oxygen and not to forget ATP (chemically bound energy; molecule). Proportionally to development of photosynthetic organs, the roots will start increasing in surface. Roots thus form, depending on the leaves surface for water and nutrient uptake, while the improved nutrient supply in turn allows the leaves to expand their biomass and thus photosynthetic activity. It all happens in circles.

The important connection at that point is: the more photosynthetically active organs a plant has, the more nutrients can also be converted into chemically bound energy (ATP), that can be used by the plant within cellular respiration. Roots and leaves resonate with each other in their growth process and are mutually dependent on each other, and it is more of a cooperation than a competition. Roots depend on leaves and leaves depend on roots.

The EC value of young plants is low because most of the energy in form of ATP is converted fairly directly for numerous growth and cell division processes. Storage of higher molecules and nutrients happens only marginally at the point, for example, to strengthen the shoot axis with a vigorous supporting tissue.

How the needs for Nitrogen, Phosphorus and Potassium change over a plants life cycle

You may have realized that most of the commercial plant nutritions contain higher amounts of Nitrogen in the vegetative formulas and more amounts of phosphorus for flowering fertilizers ("N-P-K" - the n stands for nitrogen and the p for phosphorus. K is potassium).

Nitrogen

The prioritization of energy use in the early stages of a plants life is clearly on the production of DNA, chromosomes, nuclei - the core structure of a plant, where amino acids play a big role. This needs a lot of amino acids to translate the information saved on the DNA into real plant structures. Chloroplasts have to be built up in leaves structures for performing photosynthesis. Nitrogen plays a big role in the synthesis of both chlorophyll and amino acids and in the construction of cell walls. Amino acids for example need only nitrogen out of all minerals found in a fertilizer bottle. Super simplified spoken, nitrogen plays a big role in establishing the basic frame of a plant that's later be filled with other molecules. [7]

Phosphorus

Later on during flowering, more phosphorus is needed in a plant, when a lot of energy is put into the formation of flowers. At this stage, much moveable nitrogen is already stored in the leaves for supporting new cell growth that occurs in the flowering stages when many new cells are built up for densely stacked buds and structural elements like trichomes and complex flavonoids like terpenes. For this, plants need a lot of energy to transport complex molecules through the plants cells in an active way. To do this in a short period of time, the plant needs a lot of energy in the form of ATP, which needs phosphorus to be built up - phosphorus is the only mineral out of a fertilizer bottle, that plants need for making ATP. As in the cell respiratory process way more ATP is built up than in the photosynthesis process. So plants have to establish a lot of leaves to produce glucose first, which is then transported down to the roots, where glucose can be converted into ATP in a way much larger amount than during photosynthesis. Take this just as another reason why plants need more phosphorus in the later stages. Because just then we have the basic requirements fulfilled for synthesizing huge amounts of ATP (-> glucose).

As cell replication rates increase exponentially during the flowering phase, also more DNA has to be synthesized at this stage. For this again, only phosphorus is needed out of all minerals that can be found in a fertilizer bottle. [8]

Potassium

Potassium (K) is needed equally at all stages of a plants lifetime, because this macronutrient is primarily responsible for regulatory mechanisms of the plant, for metabolic processes and support functions. A well-known plant process that's controlled by potassium is the level of transpiration by opening and closing the stomata cells at the bottom sides of all leaves . [9]

As at the beginning of a plants life there is mostly need for photosynthesis and not that much cell transpiration,

I hope this somewhat more comprehensive excursion into the world of fertilizers or nutrients has sufficiently explained the role of different ratios of fertilizer components in different stages of a plants life.
The more photosynthetic organs are available, the more ATP can be used over time for various processes, the more nutrients can be converted, moved and stored in the plant in percentage terms. It's like building a city: At first there is need for some infrastructure that's built with a lot of concrete (nitrogen) and then there is need for a lot of daily goods (phosphorus). In both the construction and the running state of a city there is need for people who run everything (potassium). Hope this comparison kinda works for you :D.

So, what's all the sience about EC for?

Now it should be clear that the EC value alone is not that meaningful. Most important is the gradient between plant salt concentration and medium salt concentration, which can be analyzed by measuring input EC and runoff EC. With gadgets like the Bluelab Pulsemeter you can even measure the EC of the medium by sticking probes in the medium.

However, experience has shown that the chart at the beginning of the article can be used as a guidance, when you don't have the abilities to measure everything. It's easy to see, that the EC value of the plant, and analogously also the target value of the nutrient solution, increases slowly but steadily during a plants life time. It's just important to make no huge jumps from low to high EC values, because this can damage a plant heavily. But with an even increase, some gardeners even can go up to EC values of 5 and higher and still have healthy plants. For such results, all parameters during the grow should be optimized in every detail.

With this knowledge, we can now also explain, why the EC value of the solution should be higher at flowering, than in the youthful stage of the plants. When flowering, cannabis plants need more nutrients, which they transport via further transport processes, from the root to the flowers, to form them nice and lush. They are also needed for nutrient storage, biochemical processes and compaction. Nevertheless, it has to be said that in the last 3-4 weeks of a grow the EC values should begin to get lower again as the plants don't produce much structures anymore during ripening.

Young plants on the other hand, which primarily perform photosynthesis, concentrate on building structure and, due to their under-prioritization of reproduction, they are not yet really interested in carrying out more complex metabolic processes for thick flowers, or are still limited in their possibilities at the beginning due to the small leaf and root surface. So they can perfectly grow with lower EC values.

The reverse osmosis unit

As i have already mentioned at the beginning of the article, the EC value of a solution in itself, has nothing to say about the number or the percentage of relevant nutrient salts in the nutrient solution. Tap water, for example, contains dissolved sodium and chloride ions, which influence the EC value, both in the nutrient solution and after uptake in the root, but they do not perform any function in the plant and rather cause damage.
For example that overall less of the usable nutrients can be absorbed in favor of the sodium and chloride. A high EC value in the plant due to useless salts, leads to water shortage and thus to symptoms such as soft, wilting leaves, growth inhibition, etc.

To remove all salts and enrich this relatively clean solution with the desired nutrients, such as nitrogen, phosphorus and potassium in a closed system, the water for the base of a nutrient solution should be run through an reverse osmosis system first. It's better to achieve the desired EC values with the bought bottled or powdery nutrients than through substances that bring no benefit but harm to our project.

The harmful ions in red and the nutritional ions in blue. Right picture shows optimum for plants.

[1] https://de.wikipedia.org/wiki/Elektrische_Leitfähigkeit

[2] http://www.hortipendium.de/Salzgehalt

[3] https://www.spektrum.de/lexikon/biologie/osmose/48395

[4] https://www.pflanzenforschung.de/de/themen/lexikon/naehrstoff-wasseraufnahme-und-transport-347

[5] https://link.springer.com/article/10.1007/BF00010968

[6] http://nawi.naturundbildung.at/wp/?page_id=3128

[7] https://www.gardeningknowhow.com/garden-how-to/soil-fertilizers/understanding-nitrogen-requirements-for-plants.htm

[8] https://ag.umass.edu/cafe/fact-sheets/fertilizing-flower-gardens-avoid-too-much-phosphorus

[9] https://extension.umn.edu/phosphorus-and-potassium/potassium-crop-production

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by Lorenz