Guest: Dr. Clay Zimmerman, Balchem Director of Technical Services This episode is from a webinar presented by Dr. Clay Zimmerman, Director of Technical Services at Balchem. To view the full webinar and access the slides referenced during this podcast, visit balchem.com/realscience and scroll down to the webinar presented on December 12th, 2023.
This episode is from a webinar presented by Dr. Clay Zimmerman, Director of Technical Services at Balchem. To view the full webinar and access the slides referenced during this podcast, visit balchem.com/realscience and scroll down to the webinar presented on December 12th, 2023.
Dr. Zimmerman begins with an overview of Balchem’s microencapsulation technologies in both human nutrition and health and animal nutrition and health businesses. (0:31)
Encapsulation is a generic term, and huge differences can exist between products that protect the same compound. Balchem’s microencapsulation technology consists of packaging a substance in a lipid capsule for protection. Encapsulates can differ in design, technology, and performance. When it comes to performance in ruminant encapsulates, stability in feed mixing and TMRs and animal performance are evaluated. (6:50)
Lipid encapsulation usually comes in one of two forms, a matrix encapsulation or a true encapsulation. A good analogy for matrix encapsulation is chocolate chip cookie dough, where some active compound is always at the surface. In the rumen, this leads to reduced protection and stability. True encapsulation, often called single-layer or multiple-layer encapsulation, is analogous to an m&m where there is no active compound at the surface, and this leads to greater protection and stability in the rumen. (12:00)
So why do we encapsulate nutrients for ruminants? In general, for targeted delivery within the gastrointestinal tract of the animal because rumen fermentation often results in massive breakdown of most of these important compounds. For example, choline chloride is almost completely degraded in the rumen. (18:30)
When developing or improving rumen-protected products for nutrients such as choline chloride, methionine, lysine, or niacin, the primary goal is to protect them as much as possible from ruminal degradation while achieving post-ruminal absorption. Once prototypes have good ruminal stability and good intestinal release, the next step is feed and mixing stability. Dr. Zimmerman goes on to showcase different research techniques for evaluating encapsulates in these three areas as well as in animal performance. (20:39)
In summary, there are many differences in encapsulated products for dairy cows, due to the design of products; types, amount, and composition of coatings; manufacturing differences; and differences in nutrient content, bioavailability, and feed stability. True encapsulates, or multi-layered coating products, are preferred for ruminant applications due to their higher levels of ruminant and feed stability. Four really important features of a good ruminant encapsulate are good ruminal stability, good nutrient bioavailability, feed and TMR stability, and ultimately biological performance. (47:05)
Dr. Zimmerman then answers questions from the webinar audiences about in vitro techniques and bioavailability, coating ingredients, the importance of base diet for rumen fluid donors in in vitro techniques, variation in products from in vitro to in vivo results, how long it takes to develop a new encapsulated product (Balchem spends years and even decades researching before a product release), and why nutrient contents differ so much in similar encapsulated products on the market. (49:58)
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Speaker 1 (00:00:08):
The following podcast is taken from a webinar presented by Dr. Clay Zimmerman, director of Technical Services at Balchem, to review full webinar and access the slides referenced during this podcast. Visit balchem.com/realscience and scroll down to the webinar titled, not All Room Protected Products are Created Equal Presented on December 12th, 2023.
Dr. Clay Zimmerman (00:00:31):
Yeah, it's quite, quite a pleasure to be here today with, with our audience to present. Not all rumor protective products are created equal, so we don't want this to be a commercial today, but thought it, we thought it would be helpful to start off maybe just giving a little background about, about Biochem and, and our expertise in the mic in the area of Microencapsulation. So, biochem Corporation, we're a publicly traded company. The company was founded in 1967. So, you know, we've been around for 56 years now, and we're basic in micro encapsulation technology. My understanding is that really almost from the start when the company was founded in 1967, we were a micro encapsulating product. So we have many, many decades of expertise in this area. The company's broken up into three core business units. We have a human nutrition and health business and animal nutrition and health business that Scott and I are involved in, and especially products business.
Dr. Clay Zimmerman (00:01:42):
We also have a very large internal r and d group in the company. We work across seven technology centers across both the US and Europe from an r and d standpoint. So we have a lot of r and d resources that are committed to micro encapsulation and micro encapsulates are heavily used in both our human nutrition and health businesses and our animal nutrition and health businesses. So, the last time I asked this question a few years ago across our company portfolio, mainly the, you know, the human nutrition and health businesses and animal nutrition and health businesses, we sell over 200 different encapsulated products. So its microencapsulation is very heavily used within the company.
Speaker 1 (00:02:48):
New research is changing everything we thought we knew about choline's impact on the cow and her calf and top scientists have a lot to say about it. They're presenting new research that supports choline as a required nutrient to optimize milk production choline as a required nutrient to support a healthy transition choline as a required nutrient to improve calf health and growth, and choline as a required nutrient to increase colostrum quantity. This new research is solidifying choline's role as a required nutrient for essentially every cow, regardless of health status, milk production level, or body condition score. Learn more about the science that is changing the game and the choline source that is making it happen. ReaShure Precision Release Choline from Balchem, visit balchem.com/scientists say to learn more.
Dr. Clay Zimmerman (00:03:52):
So, you know, as far as application of encapsulates in our human nutrition and health business, you can see some different examples here. And, and uses for encapsulation in our human businesses would be you know, first of all to help control, release and maintain the functionality of ingredients in products. It can also help improve stability and handling characteristics and in, and increase viable shelf life. So you can see there are some different applications of microencapsulation in the bakery businesses, meat businesses and confectionery businesses. So there are lots of different applications for micro encapsulates in our human businesses, as well as certainly some nutrient protection technology as well for human applications.
Dr. Clay Zimmerman (00:04:53):
As far as our animal nutrition and health business we do, we do produce some micro encapsulates for their monogastric species. We started a a pet Sure business that business started, I believe about seven years ago, and there are some different encapsulated products used there. We have pet share pH control systems that are utilized in the pet food industry, pet she porosity and texture and pet share structure and forming which, which all utilize encapsulates in, in the monogastric species. And we do some encapsulation also in Europe encapsulating tannins there. The focus of the presentation today is micro encapsulates for ruminants. So our products there, our primary products for ruminants would be ReaShure. That's our flagship product. That was our first foray as a company into the animal space in the late 1990s.
Dr. Clay Zimmerman (00:06:05):
So, ReaShure pro precision release choline. We have a newer version of ReaShure called ReaShure XC that we use in Europe currently. Nisu is encapsulated niacin. Amur XM is encapsulated methionine amur is encapsulated lysine. And the, and the last product would be nitris, that's encapsulated urea. Nitris acts a little differently than the other products, and I'll, I'll explain that a little bit later. As far as products in the ruminant portfolio, so encapsulation technology, this, this is a quote from a, an article that appeared in in Feedstuffs in 2011. Not all encapsulated products are created equally. Consequently, there are vast differences in their efficiency and stability. And that's certainly true. And that's, that's really, you know, what the focus of the presentation today is, is, is to highlight some of these differences. And what, what can lead to differences in encapsulates? So encapsulation, it's a, it's a very generic term. Huge differences can exist between products that protect the same compound, even what causes the variation in encapsulation performance. So there are a number of factors here. One, certainly, is the starting form and mode of inclusion of the active ingredients. The coating technology can vary, and manufacturing processes can vary.
Dr. Clay Zimmerman (00:07:54):
So again, micro encapsulation, it's, it's one of our core businesses is protection technology. And, and it's shown here in, in this, in this diagram, essentially when we're micro encapsulating, we're taking a substance or substances and we are protecting it in our case, in layers of lipids with our micro encapsulation. This is just a picture under the microscope of an encapsulate. This is a cross section here. So you can see the active compound in the middle and the lipid coating on the outside of the product. So encapsulates can vary in a number of different ways. They certainly can vary in design of the product and the technology that's being used to do the encapsulation, and they certainly can vary in performance performance. So there really are a couple of different things that we look at from a performance stand standpoint in ruminant encapsulates. One is stability in feed mixing, and in a total mixed ration, how well, how well do these encapsulates withstand mixing and sitting in a total mixed ration. And ultimately it's animal performance that matters with the product. How does the product actually perform on the farm?
Dr. Clay Zimmerman (00:09:30):
So, getting a little more granular, what are some of the differences in encapsulates? So first of all, it could be both starting form of the active compound and the inclusion rate of the nutrient. So for example, if we look at room protected methionine products, there are products on the market that will vary anywhere from 15% DL methionine up to 80, 85% DL methionine. So, so the act the activity rate of encapsulates can vary greatly. That's true across, you know, all of these different compounds utilized in ruminants. The coating systems utilized can vary dramatically. The composition of the coatings utilized can vary greatly and the manufacturing processes themselves can vary. So there are a multitude of factors that can lead to these differences in encapsulated products.
Dr. Clay Zimmerman (00:10:41):
If you look at encapsulates across the different industries that are out there, they're they, these encapsulates can come in all different shapes and sizes. There are a lot of different types of encapsulation that are used across industries, and they will lead to very different product characteristics. In general, when we're looking at agricultural applications, the two primary types of technology that are utilized with encapsulation are the true what we'd call true single or multiple coatings. So these first two examples here, or what we would call a matrix particle, which is shown here.
Dr. Clay Zimmerman (00:11:30):
So this is just a picture, this is just a, you know, different different products that are on the market. These aren't all biochem products that are in this picture, but you can see they do, they literally do come in all shapes and sizes and, and vary across, you know, across the spectrum here. And this can lead to very different performance, both in the feed and in the animal. So when it comes to lipid encapsulation so the, the, the biochem encapsulates are, are, are lipid encapsulates. These are the two primary types of encapsulation that are utilized in the industry. So the, this, this diagram on the left is showing a matrix encapsulation. Another term that's sometimes used for this would be spray cold and spray cold or spray chilled encapsulates soots. And the analogy that we use there would be chocolate chip cookie dough.
Dr. Clay Zimmerman (00:12:36):
So if we were, if we were encapsulating choline chloride, for instance, and a matrix encapsulation, essentially what we would do is we would, we would mix choline chloride with our lipids that we're using to encapsulate, and we would form particles from that that would look like this. With a matrix encapsulation, there's always some, some of the active compound at the surface, just the way it, the way it's mixed. In general, you know, when it comes to ruminant applications, a matrix encapsulate will have reduced ruminal protection instability. And we'll show you some examples of that a little bit later. The diagram on the right is showing a true encapsulation. So these are also known either as single layer coatings or multi-layer coatings. The example we would use here would be an m and m where essentially we're, we're, we're, we're forming a core that we are protecting in either single or multiple layers of lipids that are fully encapsulating the product.
Dr. Clay Zimmerman (00:13:54):
So in general, with a true end cap, there's no compound at the surface of the product. It is completely enveloped in layers of lipid coating in this case, which result in higher ruminal protection and stability in general. So for ruminant applications, true encapsulates are, would be the preferred application for, for mo for most ruminant applications. So this is a time lapse photograph that compares a true, a true encapsulate on the top versus a matrix encapsulate. So these are two rum protected lysine products. The flask on the left is the true encapsulate, and this is just sitting in room temperature water. So you can imagine what would happen in the room and would be more severe than this. But this is, this is just showing you, you know, through TimeLapse pictures, what happens with a true end cap on the left and a matrix end cap on the right. So you can see it times zero as soon as they're put into water, the products look different in the flask. After 30 minutes sitting in the water, you can see the product on the right is starting, the water's starting to take on some color. That color is from the release of lysine hydrochloride into the water at two hours. You can see even more color and more difference in the product on the right. And after four hours, there is quite a bit of color and release of the lysine hydrochloride into the water.
Dr. Clay Zimmerman (00:15:42):
So this is a very simple test just, just to show what, what happens with some of these products just sitting in water again, what's happening in the room and would would be, would be even greater release than this. Another characteristic that some, some encapsulates have would be freeze thaw stability. Because of all of the human food applications that we have as a company, all of our products are freeze thaw stable. So these are actually two examples here on the screen. These are products that are not freeze thaw stable. So you can see these products are both encapsulated. These are under the microscope, so the product on the left and product on the right, you can see they appear to be intact before freezing, after freezing. You can see how these cracks or fissures start to form and these products and that, that's compromising the efficacy of the product at that point.
Dr. Clay Zimmerman (00:16:51):
So if we were to test these products for ruminal protection so the product this first product is a freeze-thaw stable product. So this product before freezing is 90% room and stable at eight hours after freezing, it's still 90% room and stable. The product on the right is not freeze thaw stable. So this product before freezing is 80% room and stable. After freezing, it's less than 40% room and stable. So it loses over half of its room instability after it goes through a freeze-thaw cycle in that case. So you're, you will lose product efficacy in that case.
Dr. Clay Zimmerman (00:17:39):
This is just a picture of a, of, of a freeze thaw stable product on the left versus one on the right. This is our encapsulated urea product, nitro. Sure on the left, this is under a microscope and there's dye applied to this. There's a yellow dye. You can see the nitro. None of the dye is absorbed by the product. This product on the right is not free, is not freeze thaw stable. You can see the fissures in the product on the right when it's subjected to dye under the microscope, the dye starts to go into the end cap so that, that, that product's compromised because of the cracks in the absolute.
Dr. Clay Zimmerman (00:18:30):
So why do we encapsulate nutrients for ruminants? So in general, it's for targeted delivery within the gastrointestinal tract of the animal. And ru fermentation often results in massive breakdown of most of these important compounds. So choline choline is very extensively degraded in the rumen by the ru microbes. It's 98% plus rumen degraded. Niacin is 94% ru degraded. And the, and, and amino acids are very extensively degraded as well. Most of them 90 plus percent degraded in the rumen. So many of these nutrients need to be protected from ruminal de degradation for effective absorption by the cow post ole.
Dr. Clay Zimmerman (00:19:23):
So this was some classic research. This was done in Rich Herman's lab at the University of Maryland. This was published in the, in nine, in the late 1980s. And what he, what they did here was this this control diet had 24 grams per cow per day of choline chloride intake. So they measured intake of choline chloride and how much was showing up at the duodenum. So 24 grams of a choline chloride intake, 1.2 grams appear at the duodenum. They increased this to 178 grams of choline chloride intake. And essentially no more of this showed up at the di it was extensively degraded in the rumen. They added an extra 300 grams of choline chloride to the diet, and only one more gram of that appeared at the duodenum. So just proof that choline chloride is almost completely degraded in the rumen and, and, and needs to be encapsulated to, to perform in, in a ruminant.
Dr. Clay Zimmerman (00:20:39):
This diagram shows the characteristics that are needed in a, in a quality room and protected product. So this is basically the steps that we go through when we're either developing brand new room and protected products, or we're improving existing products. The one exception to this is nitric. We haven't encapsulated urea products. We want that product to be released in the rumen just gradually over time. But when we're protecting nutrients such as choline, chloride, methionine, lysine, niacin, et cetera, we're trying to protect them as much as we can from ruminal degradation. And we want post ruminal absorption. So the first characteristic we look at is room stability. So we want as good of a ruminal stability as we can get. If we have extensive ruminal stability with a product, we sort of lose the game from the start. So we're trying to get as much product past the room and as possible intact. Second, the second part we look at is intestinal release. So, it's a balance between intestinal release and room stability. So we want good ruminal stability, but as much intestinal release as possible from the product. Once we develop prototypes that have good ruminal stability and good intestinal release, the next step we go through is feed and mixing stability. So we go through a protocol of feed mixing that we'll discuss a little bit later as well as TMR stability.
Dr. Clay Zimmerman (00:22:34):
If a product that we're developing is not feed and mix stable, we go back to the drawing board and work on prototypes that are feum mix stable. And then ultimately we wanna be able to show a biological response in the cow. We do, we wanna do the research to prove that the product actually works on the farm. In your situations, if any of, if any of these functions are compromised, the product may fail to deliver as expected on the farm. So that there are many different types of testing that can be used for evaluating encapsulated products. The first ones are in vitro techniques. So these would be different lab techniques that can be utilized to evaluate encapsulated products. The second one would be in situ techniques in the animal, so looking at ruminal stability.
Dr. Clay Zimmerman (00:23:33):
And then finally in vivo or experiments that are done in the animal to show efficacy actually in the animal. So let's talk first about ruminal stability. So I want to talk a little bit about in vitro techniques here. So this is, there are many different in vitro techniques that are utilized certainly globally to, to look at different products as well as different encapsulates that are out there. The procedure that I'm going to show data from here is a product or a procedure called the multi-step protein evaluation or the MSPE method. This was an in vitro method that was developed in Mike van Dr. Mike Van Berg's lab at Cornell University. So all of these MSPE slides I'm going to show you, are utilizing their technique that this was published back in, in 2013. And the testing that was done here was done at Cumberland Valley Analytical Services. So this is just showing some different rumor protected lysine products as far as ruminal degradability. And you can see these products with this technique. They varied in ruminal degradability anywhere from 13% up to 29% using, utilizing this in vitro technique.
Dr. Clay Zimmerman (00:25:00):
This is these are some different rumor protected lysine products, but this is utilizing intu techniques. So ruminal release actually intu in animals. This study was actually published in 2016 in the Journal of Dairy Science. And you can see these products vary greatly in, in SIT two releases. So there, there was a product here that at six hours only had about an 11% in SIT two release, whereas another one had over 90% release and they measured release out through 12 hours in situ two. So much greater differences showing here with when we look at actual in situ two releases in the cow this is the MSPE technique, looking at some different room protected choline sources that are available. There's tremendous variation in ruminal degrade in the degradation of these products. So here's a product that with 11% al Degradability with that technique, and there actually are products that are a hundred percent ruminal release with this product. So a product like this, you would question why it is even encapsulated with 100% ruminal release.
Dr. Clay Zimmerman (00:26:29):
And then this is in situ two data from different encapsulated choline products that are there. It actually looks, the INCI two data actually looks very similar to the MSPE data on these products. So this is showing actual ruminal stability here. So here are a few product. Here's a product that at eight hours only has 11% ruminal release, it's 89% stable at eight hours. Here's another product that's essentially a hundred percent released at, at eight hours in situ two. So again, there's a widespread ruminal stability in situ with these encapsulated choline products that are on the market.
Dr. Clay Zimmerman (00:27:18):
And again, I just wanna remind you again, I showed this slide earlier just the water release, again, a true end cap versus a matrix end cap on the right. So these, these room temperature water tests, they can, they can actually be an effective means of showing what I would call a very poor quality encapsulation from, from a, from a ruminal stability standpoint. The second characteristic we're looking for, again, is intestinal release or bioavailability. So again, utilizing this MSPE technique we can look at ruminal or intestinal digestibility here. So to orient you to these graphs, I'm gonna show a few of these. This dark blue rectangle at the bottom. This is showing the ru degradable, the ru degradability of these different products. These are rum protective methionine products. So the ruminal degradability, again, varies anywhere from about 7% to about 15%.
Dr. Clay Zimmerman (00:28:33):
Utilizing this MSPE technique across these products, this light blue rectangle at the top, this will be total tract undigested nitrogen. So this is actually bypassing, in theory, bypassing digestion in, in the cow would appear in the feces. This gray box is what we want to focus on. This will be the intestinally digestible nitrogen. And you can see there's quite a variation in products here. So there's a few products here that are about 80% intestinally digestible by this technique. And one here that's only 3% has very poor intestinal digestibility utilizing this in vitro technique. If we were to look at rumen protected lysine products the intestinal digestible nitrogen varies anywhere from a high of 76% down to 42% utilizing this technique.
Dr. Clay Zimmerman (00:29:41):
So now we're gonna move into some InVivo techniques of bioavailability. I'm, and I'm gonna focus on amino acids here. So this first technique is so this is for amino chiral ru protected lysine product here. This is some work that was presented back in 2009. This is utilizing a plasma by availability technique and looking at different amaal doses of lysine hydrochloride infused into these cows, and then feeding this product to calculate a bioavailability, looking at plasma measurements of lysine utilizing this technique. This product had a 62% bio lysine bioavailability utilizing this technique. We tend not to utilize this technique much anymore. With our products, there's a lot of variability of animal to animal variability, variability, and this technique. We've looked at products that within an individual product will show by availability anywhere from a zero bioavailability up to 60 to 70% by availability, the same product in different cows.
Dr. Clay Zimmerman (00:31:08):
So there's a lot of, we have seen a lot of cow to cow variability, variability in this technique. So there are some other techniques that have been developed that are utilized to look at in vivo meth nutrient bioavailability. This particular technique is a solen meth solen methionine technique. It's an InVivo method. The products are actually fed to the cows. And essentially what you're doing with this technique is you're feeding a selen, you're feeding selenium yeast to the cow in the diet at a constant dose. And that serves as a tracer of methionine in the milk. This is a published technique. It was published back in 2009 in the Journal of Dairy Science by Bill Weiss and Norm St. Pierre out of Ohio State University. We really like this technique. Unfortunately, it only works for methionine.
Dr. Clay Zimmerman (00:32:16):
It's specific to methionine, but essentially, you can test the bioavailability of removal protected methionine product by utilizing this technique. You're measuring how much methionine from those products appears in the milk protein of the cow. These just show the results of one of these trials. This is comparing amino su XM to another product in the market. Essentially, what this is saying is these two products are equal and methionine by availability with this technique. And if we double the dose of the product twice as much is showing up in the milk protein as would be expected. There's another technique that's being utilized. This, the, there's some different stable isotope techniques that are utilized there. This is a technique that has been published out of Mark Hannigan's lab at Virginia Tech. Really looking at plasma appearance and bioavailability.
Dr. Clay Zimmerman (00:33:24):
This technique can be utilized essentially for any nutrient fed amino acids, potentially choline any other nutrient. And utilizing this technique this particular product had a 55% methionine by availability. So utilizing two different in vivo methodologies, the milk solen methionine technique, and the stable isotope technique we get equal methionine by availability here. So this slide's really just to show you how, you know, how these numbers get used then and the animal nutrition models. Then. So this particular product is 70% methionine. Again, these products can vary greatly in methionine content. Th this one has to be 70%, it's 80% room and bypass, 68% intestinal avail availability. So the methionine by availability is 54 and a quarter percent. That's simply your room and bypass times your intestinal availability is methionine by availability. The number that matters in the end as a user is metabolized on methionine. And that's simply the nutrient payload. The 70% times methionine bioavailability, 70% times 54, and a quarter percent methionine bioavailability is 38% metabolizable methionine. So if you fed 10 grams of this product to a cow, you would expect metabolizable methionine to go up by 3.8 grams in that diet.
Dr. Clay Zimmerman (00:35:13):
The next thing we look at is feed and mixing stability. We utilize mixer studies to do this. So this just talks a little bit about the technique, technique that we use internally. We do this on a small scale. We mix 100 pound or 45 kilo batches of different products. We've used this across a variety of different encapsulated products to look at, at mixed stability in a mineral mix. In our mineral, we use sodium bicarbonate, limestone, magnesium oxide, and a little bit of soybean oil for dust control. 'cause The mineral can be quite dusty. And the recipe would that we use is anywhere from 90 to 98% mineral mix and two to 10% of the end caps, depending on the inclusion rate of these encapsulates. And we mix these products for three minutes. This would be the most, really, the most severe thing you can do from a feed mixing standpoint would be to mix these with a mineral mix.
Dr. Clay Zimmerman (00:36:23):
This is a picture of the lab scale mixer that we use. It's a ribbon paddle mixer. This is a picture with a batch loaded in there. So we've got the mineral on the bottom. This is an encapsulation on the top. This is not a biochem encapsulate on top here. This would be a competitive product. So this, this is what it looks like as we load the mixer. And then after we've mixed the product, these are the results of mixed stability. This is with some different RP choline products. So again, this is showing stability in, in this graph. So this, the products here in red, so the solid line is the unmixed product. So what was stability prior to mixing, and then what was stability after it's mixed in the mineral. So this product, this is reassuring, in this case, very stable, both before mixing and after mixing in a mineral mix.
Dr. Clay Zimmerman (00:37:25):
This blue product here this will be a competitive product. It's not very room and stable prior to mixing. And the mixing didn't really damage it anymore. This product in the gray was about 70% or about 30% room and stable. Prior to mixing, after mixing in a mineral mix, it actually lost about 20 percentage units in room stability. So that product was damaged by mineral mixing at three minutes in a mineral mix. So that, so there are differences there as well. This is some tests and we've done with ReaShure, just, you know, mixing times. Our recommendation is to keep mixing times to four minutes or less inter mineral mix if we want optimal product performance. So our general re-mixing recommendations for encapsulated products, and this will really be true of all encapsulates, we recommend mineral mixing times at four minutes or less to minimize excessive mix mixing and unnecessary abrasion that can occur.
Dr. Clay Zimmerman (00:38:42):
We also recommend encapsulate adding encapsulate products as late into the mixing process as possible to minimize the total mixing time of the encapsulates in that mix. If you have at least 50% non mineral ingredients in the formula, it greatly reduces potential abrasion of encapsulated nutrients and keep encapsulated products dry in sealed bags. A lot of these products are hydroscopic, so we recommend storing encapsulated products below 120 degrees Fahrenheit or 50 degrees Celsius. Recommend a store. Storage temperatures for end caps are 50 to 90 degrees Fahrenheit, or 10 to 32 degrees Celsius.
Dr. Clay Zimmerman (00:39:35):
Finally, we wanna look at TMR stability. So there was a PA that a paper published, I referenced this one in an earlier slide. This was published in the Journal of Dairy Science back in 2016. This work was done at the, at the minor institute in New York State. They were looking at basically all of the different commercial rumor protected lysing products that were on the market at the time. This work was actually done over 10 years ago, but it was published in 2016. I will tell you, some of these products are not available commercially in the market anymore, and they measured TMR stability at two different dry matter levels of the TMR. So these solid lines in this graph were with the lower dry matter TMR, essentially 40% dry matter, 60% moisture in the TMR. The dashed lines were the higher dry matter TMR about 52% dry matter or 48% moisture.
Dr. Clay Zimmerman (00:40:43):
And you can see there's a lot of variation in TMR stability. So what essentially what they did was they mixed these products into the TMR of the two different dry matter contents. And they let them sit anywhere from zero to 24 hours. So 0 6 18 and 24 hours sitting in ATM R. All they're doing is measuring how much of the lysine was released from the product into the TMR. So the product's losing lysine to the TMR, that they're not measuring ruminal stability, they're just measuring how much lysine was lost to the TMR which means it would degrade in the rumen. So if we were to look at six hours, for instance, so this is, if you were feeding a TMR four times a day, these products varied anywhere from essentially no release in the TMR to about 60%, a 24 hour time point that would, if you're, these cows are being fed once a day. Again, some of these products had essentially no release to the TMR and others were over 50% released. So pretty big differences in TMR stability of these rumor protected lysine products.
Dr. Clay Zimmerman (00:42:04):
This is some work that we've done utilizing a similar technique, looking at different rumor protected choline products that are on the market. So this is looking at stability in this case. So this is looking at 0 6, 12, and 24 hour time points. So cows that would be fed either four, two, or one time a day and looking at TMR stability. So how much of the choline in these products is just being lost to the TMR sitting in it. So you can see two of these products have very good TMR stability essentially, you know, almost no loss in this top product. That'd be a ReaShure XC product. This is ReaShure is losing very little to the TMR. Some of these products are losing almost all of their choline payload to the TMR just sitting there. So these products are losing 70% plus at six hours over 80% and essentially a hundred percent by 24 hours. So some pretty major differences in TMR stability.
Dr. Clay Zimmerman (00:43:18):
And then ultimately we want to measure biological responses in the cow. So we'll finish up with a few slides here. So rumor protected lysine, this was some of our original amino sure l research that was presented back in 2009. So when we fed either 30 or 60 grams per cal per day of amino, sure l compared it to a controlled diet that would've been lysine deficient. We saw very nice milk production responses here. So, so increases in dry matter intake and and, and five pound increases in milk yield in this case. So demonstrating the, the product actually works in the animal with increased dry matter intake and milk yield. This is some work with ru protective methionine looking at a couple different meth ru protective methionine products here versus a control diet. And we saw the expected response here. We saw an increase in milk protein percentage, significant increase versus a control diet with both of these hormone protective methionine products.
Dr. Clay Zimmerman (00:44:27):
That would be a typical response that we would expect to see increased milk protein percent and ultimately increased milk protein yield with RU protected methionine. And of course, there's a lot of published research out there on the benefits of ReaShure, our rumor protected choline product in transition cows. And the benefits really carry through aft even after supplementation of the product during the transition period. This first study was published back in 2018 in the Journal of Dairy Science. These cows that were fed ReaShure three weeks prepartum, or three weeks postpartum, exhibited a 4.6 pound increased milk yield actually throughout lactation through 40 through 40 weeks of lactation. This is a second study outta the University of Florida, published in 2020 showing, and this was through 15 weeks of lactation. Again, the products only fed three weeks prior to having it to three weeks post calving, and there was over a five pound energy corrected milk response out through 15 weeks of lactation and some more recent research out of Barry Bradford's lab at Michigan State University showing seven pound milk responses through 12 weeks of lactation when ReaShure was supplemented.
Dr. Clay Zimmerman (00:45:59):
Three weeks, again, three weeks prior to having to three weeks post.
Dr. Clay Zimmerman (00:46:07):
And this is just showing from this was from the ars A d Meta-analysis on room protected choline products. This was published in Journal of Dairy Science back in 2020, just showing the consistent responses to supplemental room protected choline during the transition period. And, and two newer studies in higher producing cows. This is the Michigan State work I was just showing and work out of Heather White's lab at the University of Wisconsin that was published this year as well. So we continue to see these consistent milk responses in cows that transition, cows that are supplemented with hormone protected choline, even at very high production levels.
Dr. Clay Zimmerman (00:47:05):
Scott Sorrell (00:48:51):So, to summarize, we'll wrap up here. There are many differences in encapsulate products for dairy cows, and the differences are due to design of products. There, there are differences. Now, these products are designed, there are different types of coatings amount and composition of coatings. I can tell you that all of our rumen and all of our rumen encapsulated products, they vary in they vary in amount composition of the coatings. It all depends on what we're trying to accomplish with that nutrient in the cow. And where we're trying to deliver product to the cow.
Dr. Clay Zimmerman (00:47:46):
There are manufacturing differences that account for differences. They different nutrient content. Again again, I'll go back to the methionine example where, you know, methionine content can vary anywhere from 15 to 85%. And these products' bioavailability certainly differs between products. And you can see there are differences in feed stability as well. True encapsulates, or the multi-layered coating coated products are preferred for ruminant applications due to their higher levels of ruminant and feed stability. And there are four really important features of a good ruminant encapsulate good ruminal stability, good nutrient bioavailability feed, and TMR stability and ultimately biological performance. We want these products to perform on farm in your cows. Thank you.
Scott Sorrell
All right, Clay, thank you very much for that presentation. Before we get started answering your questions, we'd like to share a brief video, then we'll be right back to answer the questions submitted during today's presentation.
Speaker 1 (00:49:10):
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Scott Sorrell (00:49:58):
Alright, it's a reminder, you can still submit questions through the q and a tab at the top of your screen. Clay, first question comes in from David. He's he's commenting that he also likes the MSPE assay, but he's wondering if there are any others that you would recommend
Dr. Clay Zimmerman (00:50:15):
It's a great question. So ultimately it's the performance in the animal that matters, right? There are certainly benefits to, in vitro techniques, it's very important in the product development process, right, to help speed that up. So we utilize in vitro techniques as a screening tool on the front end during, you know, during product development. We certainly can't take the time to do all of the animal testing that would be required during a product development process. It would take many, many years to actually run all these products through animals. So the MSPE is certainly here in the us. That's the one that we prefer. There are a lot of other techniques that are out there. We've done work with different techniques. None of them, what I will tell you is there still is work lacking really aligning these in vitro techniques to what is happening in animals, especially across all these different activities that are out there with encapsulates.
Dr. Clay Zimmerman (00:51:33):
So, you know, when it comes to these encapsulated products, again, you know, they vary a lot in technology and the actives change, right? Are we talking about methionine, lysine, choline, you know, or other things. So I would say there still is a, there still is work to be done really correlating actual performance in vivo in the animal to these different in vitro techniques that are out there. It's easier to do from the room and stability standpoint, but, but ultimately when you're looking at, you know, bioavailability, I think there's still work to be done there. So it's a great question.
Scott Sorrell (00:52:20):
Yeah, Clay, we've seen people use the in vitro assays to kind of claim that's the bioavailability of their product. And so I know you kind of answered that question already, but maybe could you kind of pointedly answer that? Is that the appropriate way to measure bioavailability? And how reliable are those numbers?
Dr. Clay Zimmerman (00:52:42):
I would, I'd say again, it's a starting point. It's, it's, it's missing, it's missing some of the steps, right? These in vitro techniques, they're not taking feed mixing and feed stability into account, right? So, you know, you want to, to know how our product's really gonna perform on the farm. You have to go through the whole process. These encapsulates have to withstand mixing in the feed mill. Think about all the steps that an encapsulate will go through, being mixed in feed, being conveyed, going into feed bins, going into a bulk truck, being delivered to a farm and going into a TMR mixer. So there are a lot of steps between you know, start that starting end cap and ultimately the cow consuming it. So that's, you know, that, that's one point that certainly gets dismissed within in vitro techniques that you can replicate in animal studies is putting them through, you know, all the rigors that they need to withstand on the farm.
Scott Sorrell (00:53:52):
Yeah, thanks Clay. Osama is asking, he's kind of curious as to what we're using in our CO and I'm, you know, without giving away anything proprietary, is there some things that you can talk about in general to kind of reflect kind of some of the complexities, that's in developing coatings for various nutrients?
Dr. Clay Zimmerman (00:54:11):
It's a really good question. I'm, I'm going to not answer it directly. What I will tell you is, you know, within the industry, there are, I'll tell you first of all, this is not an easy process. You know, we've been at this for 56 years building encapsulates for lots of applications. We've actually been working in the animal space for about 30 years. I actually just found that out last week. I do, I know, I know we've been in the, we've been selling in the, in the in the ruminant space for, for 25 years now. But it's a very complex process. There is a lot, there's a lot of science to developing a good encapsulation and there's a lot of art as well. There are lots of variables, lots of variables take into account that you vary across all, you know, all these different substances that are out there. So our coatings, they're all lipid based, but, but the composition varies dramatically. And it's not any single, it's not any single coating that's out there. They vary by product and they're pretty complex in how they're built.
Scott Sorrell (00:55:36):
All right, thanks Clay. Eric's got a very interesting question. He says, in the in vitro screening steps, how important is the type of diet of the rumen fluid donor cows?
Dr. Clay Zimmerman (00:55:47):
Wow, that's a great question. I don't know how much it varies with end caps. It certainly would vary a lot depending on what types of products you're, you're, you are trying to evaluate. When we are doing it I would say you do have to be careful about that if you're utilizing different labs to run in situ. When we run in situ, we utilize one lab where they feed a consistent diet so that to eliminate that as a variable. But I'm sure that does make a difference if they're on a really high forage diet or a, you know, higher grained diet, for instance. I'm sure, I'm sure that, that, that could make some differences there because obviously you're, you could be altering the microflora, the microbiota that are in the rumen.
Scott Sorrell (00:56:49):
Alright, thank you. Paul's asking why do some products vary so much in results from in vitro testing versus the in vivo animal tests?
Dr. Clay Zimmerman (00:56:59):
I wish I knew the answer to that exactly. And I, I think it varies. I think it varies by, you know, the compound that we're talking about. So there are, there are certain, there are certain, there encapsulation technologies that look pretty similar in vitro to, in vivo depending on, you know, steps that are included there. So, you know, any, any in vitro technique, you know, we're doing the best we can to try to mimic in vivo conditions. I think it's a challenge across all of these different technologies that are out there and the different coatings to find one, one in vitro technique that really works across such a vast array of products that are out there. So I think that's one reason why, why we see, we see these differences. We're not, we're not able to adequately mimic all the conditions that are happening in vivo and the in vitro circumstances. I mean, there are big differences as you saw, big differences just in ruminal stability between some of the in vitro techniques and what happens to what actually happens in the cow. And it seems, it seems to vary some by the nutrient, right? If, if we looked at lysine, there were huge differences there between in vitro and in vivo. Choline looked more similar between the techniques, at least from the ruminal step. Yeah,
Scott Sorrell (00:58:44):
Larry's got an interesting question. I'm gonna be interested to see how you answer this one, but how long does it take to develop a new encapsulated product?
Dr. Clay Zimmerman (00:58:51):
That's an excellent question. A long time because, right, think, think about, think about the four steps we talked about, right? So we're working on working on ruminal stability and intestinal release, and we can, you know, we can certainly, we test that initially through, through in vitro techniques. We have to do it as a screening tool. Then we go into, you know, the feed mixing and TMR mixing stability. So all of that takes time. And then ultimately we're not putting a product into the market without doing the animal testing. So, you know, looking at doing production studies and bioavailability studies on, on the products and that, that can take years. So honestly, I would say a fast one, getting a product to market quick would probably, it's probably at least a two to three year process. We've had products we've been working on for 12 years, we just, some are hard to master to get the product right, you know, to get a good product into the market. It takes, it can take a really long time to go through all of these steps.
Scott Sorrell (01:00:14):
Yep. Great answer, clay. Finally we're past the top of the hour, so I've got time for one more question. Sarah's asking why do the active nutrient content levels vary so much in encapsulated products?
Dr. Clay Zimmerman (01:00:29):
So another really good question. Some of it has to do with the technology that's being used. So if you think about the diagram I was showing, you know, the matrix encapsulate versus a true encapsulate. So the physics of creating a matrix and capsule, you can only load so much active in there, right? So I in general, with a, with a matrix, encapsulate de depends on what your starting material is. You know, whether it's methionine or choline or, or lysine. So methionine, straight dal, methionine, choline, choline, chloride. So it's not straight choline, ion and lysines, typically lysine hydrochloride, it's not straight lysine, it's, you know, it's a lysine salt. So your matrix end caps in general, you have to have a certain amount of lipid in the matrix. So in general, they have, they have less loading of the nutrient. It's pretty difficult to get much more than a 50% nutrient loading into, into that, that type of product.
Dr. Clay Zimmerman (01:01:47):
Whereas a true end cap, you can vary the amount of coating, right? You're creating a core, a material that you're encapsulating and you could put a very thin core on it. So Nire is a good example of that. Nitro, sure, we actually want to degrade in the room and we're not trying to bypass the room with Nitro. It has the thin coating on, it's only 11% coating on that products, 89% urea in that case. So you can get a really high nutrient payload in that because of what you're trying to do with it and the fact that it's a true encapsulate. So you can vary the amount of coating. Some also, you know, some of these products may be a combination of active ingredients, like maybe there's both methionine and lysine in a product, right? So you're not gonna have as high of a loading nutrient load if it's a combination product. But it has to do with design and ultimately what will work in the animal.
Scott Sorrell (01:02:51):
Great answer, clay. And with that, we're gonna put a bow on this. On behalf of your friends here at Balchem, thank you for joining us today.
Speaker 1 (01:02:59):
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