Just as I suspected.....this is an important study (below) from the cyclodextrin mega-expert Dr. Joseph Pitha:
http://www.sciencedirect.com/science/article/pii/0378517392902816
hxxp://www.sciencedirect.com/science/article/pii/0378517392902816
"Effects of ethanol on formation of inclusion complexes of hydroxypropylcyclodextrins with testosterone or with methyl orange"
by Josef Pitha and Teruhiko Hoshinoa
National Institutes of Health, National Institute on Aging/GRC, Baltimore, MD 21224 U.S.A.
Received 21 June 1991;
revised 9 October 1991;
accepted 25 October 1991.
Available online 4 November 2002.
Abstract
Gradual additions of ethanol decreased and eventually abolished the formation of inclusion complexes of testosterone with hydroxypropylcyclodextrins in aqueous solutions. With hydroxypropyl-β-cyclodextrin this occured through two mechanisms. At low concentrations of ethanol (less than 30 percent), the solvent primarily acted as a competing guest compound; at higher concentrations the dissociation primarily occurred through non-specific solvent effects. With hydroxypropyl-γ-cyclodextrin only the dissociation through nonspecific solvent effects was observed. Surprisingly, when ethanolic solutions containing fully dissociated complexes were evaporated, the solid residues had properties characteristic of complexed species, i.e., they showed the rapid and complete dissolution characteristic of complexes prepared by freeze drying of aqueous solutions. That inclusion complexes were formed during the final stages of evaporation of ethanolic solution of components was confirmed by measurements of circular dichroic spectra of a methyl orange: hydroxypropyl-β-cyclodextrin combination. In this combination the spectra of included species were highly characteristic and were recorded both in aqueous solutions and in solid state after the evaporation of ethanolic solutions but not in concentrated ethanolic solutions.
Keywords: Cyclodextrin; Hydroxypropylcyclodextrin; Inclusion complexation; Solvent effect; Testosterone
I had read similar reports in the past of ethanolic solutions competing for entrance to the cyclodextrin cavity (along with the drug) due to the non-polar inner cavity of the cyclodextrin (CD)...normally, with a water solution of CD, the inner liphophilic cavity of the CD attracts hydrophobic molecules (such as testosterone or 25i-nbome)...the CD works by making "caging" or "trapping" these hydrophobic drugs inside them, then due to the CD's outer cone properties, allows them to dissolve in water so that they can deliver the drug to the surface of the mucosal membranes, where the CD then detaches but then transfers the drug through the mucosal membranes.
Earlier you may have noticed that I dissolved 100mg of 25i-nbome in 50ml of 95% ethanol (drinkable alcohol), and also dissolved 900mg of HPBCD in 50ml of 95% ethanol, doing this on two seperate magnetic stirrers, after 8 hours, the 50ml of 95% ethanol solution with the dissolved 25i-nbome was transferred via long 12" pipette over the course of about 20minutes (slowly) to the spinning 50ml of 95% HPBCD solution, then after that, the one remaining combined solution was allowed to stir for 12 to 24 hours.
The solution was stored in a jar in freezer where it would keep indefinately. The advantage to this, is that the 95% etoh solution will not freeze, so there is no need to "dethaw" it like you would have to do had you dissolved the HPBCD and drug in a water solution. Another advantage is that the etoh evaporates fairly fast (after about 1 hour, the blotter paper is completely dry) when applied to filter paper blotter, so long as a fan heater is kept pointed at the paper for the duration of evaporation.
Keep in mind that I also sprinkled about 5 to 10mg of additional HPBCD powder on the 1.5" x 5/16" wide filter paper blotter to cover the surface of it before I dropped the drops onto it via insulin syringe needle tip. I did this for "added insurance" that the complex would form as the etoh fully evaporated from the paper in the final stages.
In the study above we see that due to the competition of ethanol for the inner cone of the CD, we will not get a complex of 25i-nbome to the HPBCD until after the drops of solubized drug are transferred to paper blotter, and then allowed 45 minutes to 1 hour for complete evaporation of etoh from the blotter, finally leaving a complex of HPBCD to drug on the paper, this occurs only after all the etoh has evaporated,
so in essence, I have been on the right path, as I always applied the blotter after all the etoh had evaporated, when the paper was completely dry, then I would apply to upper gum, and it worked fantastic.
So in other words, the way I have been complexing has been working due solely to the HPBCD complexing to the drug as the etoh evaporates from the blotter paper, causing a complete complex in the end.
"Surprisingly, when ethanolic solutions containing fully dissociated complexes were evaporated, the solid residues had properties characteristic of complexed species, i.e., they showed the rapid and complete dissolution characteristic of complexes prepared by freeze drying of aqueous solutions. That inclusion complexes were formed during the final stages of evaporation of ethanolic solution of components was confirmed by measurements of circular dichroic spectra of a methyl orange: hydroxypropyl-β-cyclodextrin combination. In this combination the spectra of included species were highly characteristic and were recorded both in aqueous solutions and in solid state after the evaporation of ethanolic solutions but not in concentrated ethanolic solutions."
So you have a few choices when you complex HPBCD to 25i-nbome:
1. you can do it the way I did above, but remember that the HPBCD will not complex to the drug until the final stages of the ethanol has evaporated from the blotter, after the blotter is dried, then you will have HPBCD-complexed to 25i-nbome.
2. you can dissolve HPBCD powder in distilled water, then drop in your 25i-nbome drug and allow it to spin 12 hours or so, then the HPBCD will easily be complexed to the drug in solution, then just drop the appropriate amount of water drops on blotter, allow it to dry, and it's ready. This could also be used nasally (as HPBCD improves nasal absorption as well), if you add this solution to an empty metered nasal pump (such as the nasa*ort spray pum), then each spray of the nasal pump will equal approximately 0.100ml. To sterilize this solution, suck it up in a 100ml plastic syringe, screw a 0.22 micron whatman filter to the end of the syringe, and push the contents using your thumb to press on the plunger until the solution gets pushed thru the filter into a pre-alcohol washed empty metered nasal bottle. Then add say 5% (exact figure not known) of 95% etoh to the nasal bottle solution as well, to keep it from growing bacteria.
What are cyclodextrins?
Cyclodextrins are a form of linked carbohydrates. They're formed by an enzymatic synthesis that begins with starch. The enzymes, called transglycosidases, are derived from bacteria. What these enzymes do is couple the starch molecules together to form a truncated, conical, molecular structure with a hollow cavity inside.
The inside of this cyclodextrin "cone" is just about the perfect shape and volume to accommodate a steroid molecule. It's also a non-polar molecule, which means that it has some of the same properties as a fat or oil. The steroid molecule doesn't just sit inside the cone, it actually attaches to the inside of it. Also, it won't dissolve in water. However, while the inside of the cyclodextrin cone is non-polar, the outside is polar, which means that it will dissolve in water. What's the significance of all of this? When a steroid molecule and a cyclodextrin molecule hook up, they form a 1:1 complex. So, while the steroids themselves won't dissolve in water, a cyclodextrin/steroid complex will. The upshot is that steroid complexes become more absorbable through the oral mucosa.
A lot of clinical research has been published on the use of sublingual cyclodextrin complexes (SCCs) in humans. At the forefront of much of this research has been Josef Pitha of the US Department of Health and Human Services. Pitha has several patents on sublingual cyclodextrin complexes. He's also authored a journal article where he details the results of an SCC of testosterone on men. In a nutshell, Pitha found that an SCC containing 10mg of testosterone per tablet raised testosterone levels astronomically high (900% over baseline at one hour) and at two hours the levels were still elevated 485%. Compare that with another study that used regular testosterone at 20 times the dosage used in Pitha's study. Regular testosterone - not complexed with cyclodextrin - only raised testosterone around 500% at the peak.
Another study performed by Stuenkel et.al. showed that testosterone SCCs of 2.5 and 5.0mg raised testosterone levels in hypogonadal men 2341% and 4270% (absolute increases of 1765 ng/dL and 2406 ng/dL) respectively! It took an average of 20 to 30 minutes to achieve maximum blood testosterone levels, but even after eight hours post-dose, the testosterone levels were still elevated 126% for the 2.5mg dose and 195% for the 5.0mg dose. Interestingly enough, the peak levels for estradiol only increased 300% and 340% over baseline, respectively. Remarkable, considering that one usually sees estradiol levels increase proportionally with testosterone levels when other forms of administration are used (i.e. injectable esters and TU orals).
![:\](https://bluelight.org/xf/BL_Images/Orig_Emoji/wrygrin.gif "wry grin :\")
I would opt for a higher dose over this.
