World Class Bodybuilding   Forum

BulkSupplements Egg Whites International A1Supplements.com i-Supplements Strength.com VitaSprings Myprotein Beyondsupplements
Go Back   World Class Bodybuilding Forum > Anabolic Steroid Discussion > Bodybuilding Peptides and Insulin

Shoutbox
Loading...


Bodybuilding Peptides and Insulin Discuss peptide cycles, doses, how to mix HGH, IGF-1, MGF, Insulin and many other anabolic Peptides




Reply
 
Thread Tools Search this Thread Display Modes
  #21  
Old 04-10-2017, 12:32 PM
chadd77's Avatar
chadd77 chadd77 Is online now
Community Veteran

 
Join Date: Feb 2016
Posts: 623
Thanks: 149
Thanked 201 Times in 154 Posts
Rep Power: 441
chadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond repute
Quote:
Originally Posted by rocketw19 View Post
I am actually kind of seeing that with myself but only chad has actuall took the time that I can find aand do these tests. it cost money takes a lot of time and for what in the end of the day you know that's it.

but back on point yes I am seeing that with me 2.5ius had me at 350IGF and 5iu had me at 700IGF so not quite but a pretty good guess and if so chad IGF level should of been around 550-600 on 4IUs prob the lower side.
chad what was your IGF result on the ones I gave you. (I am not a source hes my friend) the black tops tested on par IGF wise with my USA serostims so I can only assume the ones I gave you to be dosed correct
In my first post I did say that my igf-1 test came back at 645 running 4iu a day on the black tops. It was my highest number in the 4 tests I've done over the last couple months. I'll be running another test in 10 days and then I think I might take a break from testing. lol Too much time and money to do it.
Reply With Quote
Get your liquid egg whites here at EGG WHITES INTERNATIONAL

  #22  
Old 04-10-2017, 12:46 PM
rocketw19's Avatar
rocketw19 rocketw19 Is Off Line
Community Veteran

 
Join Date: Feb 2006
Posts: 2,618
Thanks: 522
Thanked 574 Times in 428 Posts
Rep Power: 449
rocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond repute
Quote:
Originally Posted by chadd77 View Post
In my first post I did say that my igf-1 test came back at 645 running 4iu a day on the black tops. It was my highest number in the 4 tests I've done over the last couple months. I'll be running another test in 10 days and then I think I might take a break from testing. lol Too much time and money to do it.
don't blame you at the end of this me and you the only ones to really try and find out this info and no one gives a hoot. no one is testing IGf but me and I have done that for years no one is testing IGF on USA Gh cept me. I know what my IGF score is with USA GH I compare it to generics and its funny that the main guy we all know him its not a big mystery always has proper dosed generic GH always.

I seen in the other site they are flocking to glycos post of that's pretty good on your score but nothing else, lol not the 4 pages of what they are doing is wrong.
__________________
www.RockThisGear.com
Reply With Quote
The Following User Says Thank You to rocketw19 For This Useful Post:
chadd77 (04-10-2017)
  #23  
Old 04-10-2017, 12:52 PM
rocketw19's Avatar
rocketw19 rocketw19 Is Off Line
Community Veteran

 
Join Date: Feb 2006
Posts: 2,618
Thanks: 522
Thanked 574 Times in 428 Posts
Rep Power: 449
rocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond reputerocketw19 has a reputation beyond repute
your score is higher than mine but nothing dramatic. id say we are pretty good indicators of what your IGF score should be on proper dosed GH.

is there any board out there that is really big on info about GH? If so someone PM me someone has to be smart enough to test IGF levels on real GH so they can compare. or they prop don't care cuz they know its real and that all they are doing lol.
Im gonna call my HRT doc and see if he will give me an idea if scores are all over or the place or pretty basic avg. I know he only scripts 1iu per day and its crazy loot. is that even legal for him to do lol
__________________
www.RockThisGear.com
Reply With Quote
  #24  
Old 04-19-2017, 08:25 AM
chadd77's Avatar
chadd77 chadd77 Is online now
Community Veteran

 
Join Date: Feb 2016
Posts: 623
Thanks: 149
Thanked 201 Times in 154 Posts
Rep Power: 441
chadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond repute
Just got my blood work done this morning on the Ansomone I've been running at 4iu a day. I tested Ansomone in the past but this is from a different supplier. Ansomone is supposed to be pharm grade hgh but I have my doubts about that and it's why I'm running another test. I should get my results back in 2-3 days.
Reply With Quote
  #25  
Old 04-19-2017, 12:29 PM
MR. BMJ's Avatar
MR. BMJ MR. BMJ Is online now
Administrator

 
Join Date: Jul 2008
Posts: 9,669
Thanks: 5,398
Thanked 2,244 Times in 1,719 Posts
Rep Power: 2130
MR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond repute
We/members are getting a lot of the HGH tested over at proM/anasci. Hopefully the results come out soon, but the info has to stay on those forums and not shared elsewhere since people are donating funds to help the costs.

There was an interesting debate on how (real) HGH is assimilated by each person's body, and therefore, possibly giving different results when doing IGF/serum testing. It was a discussion between both Homonunculus and Janoshik...let me know if this link does not work for you:

http://www.professionalmuscle.com/fo...und-2-a-2.html

If it doesn't, i'll copy and paste their responses/discussion. If it does work, starting from post #23 onward is where it starts. Gives some food for thought on all this.
__________________
Reply With Quote
  #26  
Old 04-19-2017, 02:08 PM
chadd77's Avatar
chadd77 chadd77 Is online now
Community Veteran

 
Join Date: Feb 2016
Posts: 623
Thanks: 149
Thanked 201 Times in 154 Posts
Rep Power: 441
chadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond reputechadd77 has a reputation beyond repute
It's not allowing me to view that page, I even created an account. I'd be interested in seeing what they have to say.

I get that everyone is different but my tests should show which brands are testing better than others. I understand if my igf levels come back at 400 yours on the same brand and dosage could come back at 450.
Reply With Quote
  #27  
Old 04-19-2017, 03:00 PM
MR. BMJ's Avatar
MR. BMJ MR. BMJ Is online now
Administrator

 
Join Date: Jul 2008
Posts: 9,669
Thanks: 5,398
Thanked 2,244 Times in 1,719 Posts
Rep Power: 2130
MR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond repute
Buck1973:
Quote:
Mr. Jano,
How is the testin that was done last time and what you propose this time different? you sugest Sec-HPLC and RP-HPLC, No Immu?????

the SEC- HPLC test is new this time , whats the dif in the 2 HPLC tests?????

The MALDI-TOF has been done here Before. I think the costs of these are more than twice as Much as the HPLC tests wat if any benefit would this get us?

Can you post up actual Tests of all these and xplain how these are read?????
We covered alot of this in PVT . I think this would be good to see by all....

Janoshik:
Quote:
I believe that the quantitative information provided by immunoassay alone was not enough for the community.

SEC-HPLC is a separation by HPLC dependent on the molecule size - it is also used in pharmacopoeia and is good to determine amount of fragments or dimers of HGH, which is a very valuable information.

Dimers and fragments appear in pharma HGH and generics alike and from what I've seen up until now they are the major protein contaminant in both. What is also important is that they do not pose a hazard for one's health.

What does that mean? That the same impurities appear in both pharma and generics and the major difference in the amount of those - if any other protein contaminant appears, it will be possible to approximate the molecular size of it and to tell if it 'should' be there. If it's completely random protein then it might be a cause to worry.

The setup I am using has specs much better than those requested by the European Pharmacopoeia.

It had been very hard to optimize the method and the chemicals involved are scary to me, even, however, after some experimenting etc I was able to get variance of about 0.05% (!), which is much better than I had ever hoped for.

I have attached two examples of SEC analysis on pharma sample with dates and everything on them - so people can see the extremely tiny variance of the same sample even two days apart. Maybe somebody can chime in and elaborate on how exceptional it is to achieve data variance so low.

RP-HPLC is what was done before - it is separation based on polarity, much improved since the last time as well.

MALDI-TOF provides very precise information about the molecular weight of the sample and the contaminants - making it much easier to identify the unknowns. These would be conducted by a facility independent from me, I'd only be providing samples. Was able to get the price as low as the other testing methods cost, but it would not be my first choice.

Racepicks:
Quote:
Based on the above information, I am most intrigued by the SEC-HPLC because of it's stated ability to determine the amount of fragments, which I am guessing are broken Amino Acid Polypeptide chains.

I believe that Homonunculus referred to a study that mentioned those fragments may be what seperates the effectiveness of generic and Pharma. Let me see if I can get him over here to expand on this. Or at the very least, set me straight.

Janoshik:
Quote:
I'd be very happy to discuss that!

Like I mentioned - in generics I've had the possibility to test with SEC yet I have not noticed contaminants that were not in the pharmaceutical preparation. Just the amounts were vastly different, so it's possible that's the cause of the variance of ones response to the preparation.

It would be nice to get some anecdotals and correlate them with amounts of some fragment/derivative etc. Not saying it HAS to be working that way, but something interesting might be found out.

After I get some sleep I may look into some articles regarding that.

homonunculus:
Quote:
So this is all pretty complex stuff, as there are a number of fragments, two main isoforms (22kDa - "main one" that's made via recombinant DNA technology and the 20kDa - truncated version that has less impact on reducing insulin sensitivity.) Some of the fragments have biological activity and some may not.

Levels of these fragments and isoforms vary according to their clearance from the plasma, too.

Here's a nifty study proposing a way to measure immunofunctional activity (binding to antibodies) that would represent what of these proteins (iso forms and fragments) are biologically active, based on the idea that bioactivity requires dimerization of the GH receptors (two pieces of the receptor coming together d/t binding of an isoform or fragment).

https://www.ncbi.nlm.nih.gov/pubmed/8675586

Another issue is that different isoforms and fragments have different half-lives in the blood - get cleared at different rates.

https://academic.oup.com/jcem/articl...jcem.86.1.7129

So, if you take a sample that has a given mix of fragments in it and quantify these, this is a static picture of the iso form and fragment composition of the same.

In terms of effectiveness of the sample, some of those fragments (you'd have to figure this out from the literature) would be bioactive and others not. Also, sme fragments might hang around for awhile and thus have greater bioactivity, too, and this could vary depending on whether the product was administered before / after exercise, etc. (See link above.)

So, I think there's still some guesswork here as afar as bioactivity, even if one knows a lot about the fragments, etc.

Mass spec will tell us about oxidation, deamination and cleavage (1. Jiang H, Wu SL, Karger BL, and Hancock WS. Mass spectrometric analysis of innovator, counterfeit, and follow‐on recombinant human growth hormone. Biotechnology progress 25: 207-218, 2009. )

Whether or not oxidation matters to binding prob. depends on the amino acid residue affected:

1. Mulinacci F, Bell SE, Capelle MA, Gurny R, and Arvinte T. Oxidized recombinant human growth hormone that maintains conformational integrity. J Pharm Sci 100: 110-122, 2011.
2. Teh LC, Murphy LJ, Huq NL, Surus AS, Friesen HG, Lazarus L, and Chapman GE. Methionine oxidation in human growth hormone and human chorionic somatomammotropin. Effects on receptor binding and biological activities. J Biol Chem 262: 6472-6477, 1987.


I attached summary figure from this article with a rundown of the isoforms and fragments found in human blood: 1. Baumann GP. Growth hormone isoforms. Growth Hormone & IGF Research 19: 333-340, 2009. http://www.growthhormoneigfresearch.com/

.

Racepicks:
Quote:
I'll be honest, I have not read the links yet. I do have a question though. Is there a particular Lab Test that will identify the actual bio-active partial chains? If that was possible, perhaps we could determine the difference between these chains in generic, as compared to Pharma.

homonunculus:
Quote:
That's addressed in the first link

"The immunofunctional hGH assay is suitable for clinical
routine use, as characterized by its working range between
0.1-100 pg hGH/L. We have shown that endogenous GHBP
levels in physiological concentrations do not result in significant
interference with the hGH results derived from the
IFA method. By comparison of results from 128 sera measured
in both a competitive polyclonal RIA using the same
rhGH reference preparation 88/624 and the IFA, it can be
concluded that, on the average, 27% of molecular forms in
circulation give rise to a signal in the RIA, but not in the IFA.
From the design of the IFA, we conclude that these 27% of
hGH forms in circulation do not possess both binding sites
for the receptor and, therefore, would not be able to initiate
a signal transduction process at target cells, which is known
to require receptor dimerization. When comparing the results
obtained by use of the IFA and rhGH as the calibrator
to those obtained from polyclonal RIA calibrated by use of
pituitary hGH, the IFA results were, on the average, only 30%
of those from the RIA. The major contribution to this discrepancy,
however, is from the different potencies of the
calibrators. Direct comparison of RIA results obtained by use
of pituitary-derived VS. recombinant calibrator shows a decline
in hGH levels to less than 50%. The remainder of the
discrepancy between IFA and RIA results accounts for molecular
forms not recognized in the IFA, but giving a signal
in the RIA."

Again, this method presumes that GH receptor dimerization is needed for GH signal transduction, which seems to be true:

1. Brooks AJ, and Waters MJ. The growth hormone receptor: mechanism of activation and clinical implications. Nature reviews Endocrinology 6: 515-525, 2010.

The thing is - as we know from the differences in the effect of the 20kDa vs. 22 kDa form - simply binding and transducing a signal doesn't tell us WHAT signal in particular is being transduced, per se.

[Truth be told, I'm not an expert in the least in this area - I've just looked into it a bit. ]

rAJJIN:
Quote:
Now can you tell us in simple every day terms what testing method you propose?
Many have been listed in detail.


homonunculus:
Quote:
No, I can't proposed a testing method b/c this is a matter of access to said equipment, cost and what folks are willing to pay. Like I said, I'm no expert here.

This is a new bioassay that seems to be cheaper, faster, etc.:

1. Maimaiti M, Tanahashi Y, Mohri Z, and Fujieda K. Development of a bioassay system for human growth hormone determination with close correlation to immunoassay. Journal Of Clinical Laboratory Analysis 26: 328-335, 2012. EBSCOhost Login


Serum growth hormone (GH) level is measured largely through immunoassays in clinical practice. However, a few cases with bioinactive and immunoreactive GH have also been reported. We describe here a new bioassay system for GH determination using the BaF/GM cell line, which proliferates in a dose-dependent manner on hGH addition; cell proliferation was blocked by anti-hGH antibody. This bioassay had the lowest detection limit (∼0.02 ng/ml) reported thus far and the highest specificity for GH. The bioassay results were compared with those of an immunoradiometric assay across 163 patient samples in various endocrine states. A close correlation (the ratio of bioactivity/immunoreactivity was 1.04 ± 0.33, mean ± SD) was observed between bioactivity and immunoreactivity in these samples. The newly developed system is a specific, sensitive, easy, and fast bioassay system for GH determination; we consider it useful for evaluating GH bioactivity in various endocrine states.; © 2012 Wiley Periodicals, Inc.

rAJJIN:
Quote:
I think as members we ALL want the same thing. What method that is I am not really sure either.
I do appreciate your contributions! Everyone here knows you are one of the most respected guys.

homonunculus:
Quote:
There is power in understanding the science, in particular it's limitations. To want to boil things down to a singular "best" method is a presumption that there is a such a best thing, i.e., that we are all the same in terms of biological response, when this is simply not true.

An analogy that is really just as sciencey, but that most here are familiar with and thus a good comparison, is binding to the androgen receptor of a given steroid. One might try to say, "OK, the stronger the binding affinity, the better the steroid - it's as simply as that. So, we just need to measure androgen binding affinity of various samples of steroid X to figure out which is the purest / best of them."

People will argue with their own experience that the above simply does not carry over to real world gains on a gram by gram basis. Of course, the pharmacology of AAS is different than that of GH isoforms, etc., but we'd be making the same mistake if we just tried to look at sample testing in an overly simplistic way.

So actually, with my level of knowledge, you asking me what the best testing method for GH would be is like you asking me what the best method for determining what the best AAS is. It seems to me it's complicated to the extent that I couldn't (at least yet) justify my answer, at least when it comes to that kind of a very broad question.
__________________

Last edited by MR. BMJ; 04-20-2017 at 12:02 PM.
Reply With Quote
  #28  
Old 04-19-2017, 03:11 PM
MR. BMJ's Avatar
MR. BMJ MR. BMJ Is online now
Administrator

 
Join Date: Jul 2008
Posts: 9,669
Thanks: 5,398
Thanked 2,244 Times in 1,719 Posts
Rep Power: 2130
MR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond repute
janoshik:
Quote:
The articles are interesting, but talk about issues that are mostly not too relevant - as the are talking about in vivo byproducts and fragments, which are different from the ones that appear with rHGH. So those are different fragments than those appearing in the vials of rHGH.

Pharmaceutically, only 22kDa is being manufactured for human use and while other isoforms are possible to buy I had not found informations about them being readily avaiable at scale that would suggest them being used for anything other than research.

Immunoassays had moved long way from late 90s as well and are show quite lower variance than the one quoted in the article, which is great. It is possible, that the statement that immunoassays are the most precise way of HGH quantification still stands true.

We've talked about bioassays before, and I took a look in my mailbox - as I had inquired about the prices. It would come at about 1400$ per test for the material for bioassay alone [not counting the controls!!]- that is without the equipment necessary (the reader). I have inquired also about the BaF/GM cell line mentioned above.



In short:
So while the bioassay might be the best way to judge the effectiveness of the hormone DIRECTLY, it would not provide us informations about other protein related impurities. It is also expensive as hell and hard to work with.

I think, and this is my personal opinion, that the dosage and impurities are of much greater concern, if for nothing then for the rate that the issues with these appear.

Because if something has an exact (MALDI) or approximaly exact (SEC) length as HGH, same polarity (RP HPLC) as HGH, same hydrodynamic radius [ie shape] (SEC) as HGH or same binding sites as HGH (immunoassay) it can be assumed it holds the bioactivity of HGH, which would be confirmed by bioassay.




Also effectiveness of the hormone can also be judged indirectly from the effect on the IGF.


Like mentioned, there is no single test that is ultimate, so the best solution is a combination of them - and I believe the combination of RP and SEC might suit our needs the best, as both are capable of quite precise quantification and assesment of impurities as well.


Some words about common impurities:

An interesting quote regarding desamido and sulfoxide derivatives:
"Two derivatives of biosynthetic human growth hormone, a sulfoxide and a mixture of two monodesamido isomers, have been isolated and characterized. The sulfoxide derivative arises from an oxidation of Met-14. The major site of deamidation is at Asn-149 with a minor site at Asn-152. In addition, a fraction has been isolated from a sample of human growth hormone that was maintained at 40 degrees C for 2 weeks. This fraction, the isolated impurities fraction, contains the sulfoxide and the desamido forms, thereby demonstrating that these derivatives are the primary degradation products of biosynthetic human growth hormone. The sulfoxide, the desamido, and the isolated impurities fraction exhibit full biological activity."

So even if huge amounts of desamido or sulfoxide derivatives are found, it doesn't mean that the HGH is ineffective!

It can, however, imply improper storage or manufacturing conditions. It is also suggested by Hepner (2005) that such modifications can lead to immunogenic response. Thus potentially causing issues seen with generics (my speculation) and possibly health issues (according to Hepner as well).
Like I mentioned - it might be very interesting to correlate amounts of these with issues commonly seen with generics!

Regarding dimers:
"The somatogenic activity of such dimers, whether covalently linked or otherwise, appears to be around 10% of that observed for monomeric 22 kDa GH"

Regarding oxidated forms:
" It is not known whether oxidation of methionines in recombinant human growth hormone leads to functional impairment but conformational changes are unlikely as proposed by circular dichroism and 1H-NMR studies [33]. It is worth mentioning that oxidised methionines are not localised at the receptor binding site."




Oxidated, desamido, sulfoxide and dimers are all picked up and identified by RP (first three) and SEC (dimers) HPLC.

Other modifications/fragments are picked up by those tests as well, but the identification is not possible with my capabilities.

homonunculus:
Quote:
Janoshik,

Now I have questions BTW, great point about the endogenous vs. rHGH, as well. I didn't see a point in getting into that given the way things were going.>

You quoted, "The sulfoxide, the desamido, and the isolated impurities fraction exhibit full biological activity."

Was there a reference following that statement, or can you link the article that came from?...

So it sounds like, with the technology you have or are looking into, can you quantify the dimer and oxidized form content of a rHGH sample, as well as the sulfoxide, the desamido, and the isolated impurities(?)...

What % of a sample would be made up of the above, typically?...

This would get closer to a composite picture of the bioactivity of a product.

Of course, pharmacokinetics of said impurities might be unknown, so a biologically active impurity, even if found in "large" amounts, that has a a half life of 12 min. after injection might not have meaningful effect

Janoshik:
Quote:
https://www.ncbi.nlm.nih.gov/pubmed/3219192

It is an old article, I keep that in mind, however I vaguely remember the same fact from other articles I have read over the time.

It also makes sense from structural standpoint, as those modifications do not appear on the binding site and should not modify the structure of the hormone as well, so I don't have a reason to distrust that statement.


The quantification of impurities can be either direct - with use of a standard, which desamido form has commercially available. I am not sure about the others.

However, in non-research scenarios I prefer semiquantitative approach by normalisation. Either absorbtion at either 214 or 280nm or fluorescence can give us a pretty good idea about the amount of protein, even without a standard.

In the worst case, when standards are not available and precise quantification is necessary the standards can be prepared by chromatographic refinement.


A pharmacological preparation I have tested consisted of >98% HGH. Generics I have tested varied VERY widely. From about 3% of impurities related to hgh to almost 20%.


However, the mere fact that I am able to separate the components easily, means that it can be also purified on commercial scale easily, as I do not consider myself an expert - so I am not sure what causes the issues with impurities.

Maybe it's just skipping on some steps of purification because it is lengthy? Or is it improper storage? Lyophilisation? I'd love to say otherwise, but I truly do not have any idea.

janoshik:
Quote:
I am very tired (it's almost 4 am here now), so I hope I'm making some sense - I will try to provide short simple answers to some questions that seem to be frequently asked or could be frequently asked.


Is it possible for chinese to pass HGH releasing as HGH with any of the testing methodologies mentioned?

Absolutely not.


Is it possible to be 100% sure about the activity of rHGH in ones body?

Absolutely not.

Even if the rHGH is absolutely proper, it is possible that one has issues with processing 22kDa HGH. Everybody is unique.

And every test can fail, even bioassay, which has inherent problems from being biological assay. I believe it does not provide enough benefit for the price.

Inherent problems with bioassays can be generally summed into cells dying or mutating, both of them happening more often than research professionals care to admit to people funding them.


Can fragments / impurities have same effect as HGH?

Yes.

However, they are sign of improper manufacture or storage and can cause health issues.


Are the fragments found in rHGH and in vivo (in human body naturally) the same?

As far as I have found out no, they are not.



Would those fragments be picked up by some testing methods?

Yes, RP and SEC can also provide us with sortof qualitative information as well.

The fragments could be picked up as full-chain 'correct' HGH by immuno or bioassay.

MALDI can even tell us the exact bond the breakage happened, with some extrapolation.


Why the switch from advising to use immunoassay to HPLC?

My possibilities and experience with HPLC of HGH had increased immensely in the meanwhile.

I have also invested a lot money into specialised equipment.

I do now believe that the benefit of exact quantification gained from immunoassay is neglible, compared to refined chromatographic methods.

Was not HPLC used by SIMEC as well?

Yes. I actually am using a method adaptation from european pharmacopoeia that SIMEC had used as well.

Considering we have raw data from SIMEC and raw data from me I'll be happy if anybody knowledgeable took a single glance on the data to see the obvious and staggering is the difference in quality of the data and resolution.

Why is the shape of the hormone (if it fits like a key in the lock) important?
To get mg / vial and purity it is a necessary information.


If there is 4mg of the hormone in a vial and it's all shaped like letter M (hypothetical correct shape), we have 4mg and 100%.

However, if we have 2mg shaped like M and 2mg shaped like W, which might seem stupid, as it's the same shape but mirror image (simplified hypothetical situation) it might not work at all, so we have 2mg/vial and 50% purity.


I'll be going to sleep now, so I'll be looking forward to discussion and any questions tomorrow.


Good night.

homonunculus:
Quote:
Quote:
Originally Posted by janoshik View Post

https://www.ncbi.nlm.nih.gov/pubmed/3219192

It is an old article, I keep that in mind, however I vaguely remember the same fact from other articles I have read over the time
.

Hmmm... I don't access to that full paper (for free - I've put in an order for it), so no way to tell (from the abstract alone) how they determined full biological activity, but it is VERY interesting that they created the sulfoxide and desamido impurities from exposure to 40˚C for two weeks (and that this impurities were still biologically active, as they claim).

Quote:
It also makes sense from structural standpoint, as those modifications do not appear on the binding site and should not modify the structure of the hormone as well, so I don't have a reason to distrust that statement
Makes sense.

Here are some more interesting papers:

1. Mulinacci F, Bell SE, Capelle MA, Gurny R, and Arvinte T. Oxidized recombinant human growth hormone that maintains conformational integrity. J Pharm Sci 100: 110-122, 2011.
2. Mulinacci F, Poirier E, Capelle MA, Gurny R, and Arvinte T. Influence of methionine oxidation on the aggregation of recombinant human growth hormone. European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik eV 85: 42-52, 2013.
1. Jiang H, Wu SL, Karger BL, and Hancock WS. Mass spectrometric analysis of innovator, counterfeit, and follow‐on recombinant human growth hormone. Biotechnology progress 25: 207-218, 2009.

Quote:
The quantification of impurities can be either direct - with use of a standard, which desamido form has commercially available. I am not sure about the others.

However, in non-research scenarios I prefer semiquantitative approach by normalisation. Either absorbtion at either 214 or 280nm or fluorescence can give us a pretty good idea about the amount of protein, even without a standard.

In the worst case, when standards are not available and precise quantification is necessary the standards can be prepared by chromatographic refinement.
Plus, with the testing purposes here, you'll be able to compare "brand" A vs. "brand" B. Will be interesting to see then how folks impressions of those two match up.

Quote:
A pharmacological preparation I have tested consisted of >98% HGH. Generics I have tested varied VERY widely. From about 3% of impurities related to hgh to almost 20%.

However, the mere fact that I am able to separate the components easily, means that it can be also purified on commercial scale easily, as I do not consider myself an expert - so I am not sure what causes the issues with impurities.

Maybe it's just skipping on some steps of purification because it is lengthy? Or is it improper storage? Lyophilisation? I'd love to say otherwise, but I truly do not have any idea.
Well, heat is one thing per the study you mentioned above. (When I get it, I'll try to remember to report back. I would guess they discussion other known ways that impurities can come into being. I'll google around a bit when I've got more time.)

-------------

Your Q and A post a few posts up is AWESOME!!!
__________________
Reply With Quote
  #29  
Old 04-19-2017, 03:12 PM
MR. BMJ's Avatar
MR. BMJ MR. BMJ Is online now
Administrator

 
Join Date: Jul 2008
Posts: 9,669
Thanks: 5,398
Thanked 2,244 Times in 1,719 Posts
Rep Power: 2130
MR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond reputeMR. BMJ has a reputation beyond repute
There are a few other posts i'll add later on when I get home
__________________
Reply With Quote
The Following User Says Thank You to MR. BMJ For This Useful Post:
chadd77 (04-20-2017)
  #30  
Old 04-20-2017, 07:58 AM
Glycomann's Avatar
Glycomann Glycomann Is online now
Lumpy Moderator

 
Join Date: Nov 2009
Location: Chopping wood or laying pipe.
Posts: 10,454
Thanks: 1,084
Thanked 4,495 Times in 2,845 Posts
Rep Power: 1811
Glycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond reputeGlycomann has a reputation beyond repute
I've read through about 3/4 of this exchange. Mostly what they are describinb is the range of analytical and clinical tests used for GH analysis. There is some confusion about the tests. Mostly irrelevent confusion in a practical sense. One area that they are confuces revolves around fragments. They are confusing natural isoforms with fragments resulting from manufacturing. recombinant GH is generally only the 22 kDa form from my recollection. there is no 20 kDA form in the recombinant products unless some company is going outside of the norm and has produced their own vector for production.

Fragments in the manufacturing sense is consequential. These are produced either by proteases or mechanical cleavage durring the production process. Another issue that was not discused is aggregate. Aggregate forms in a lot of recombinant pharma protein products and can be immunogenic and that is not good. So if you see a bunch of high molecular weight stuff in yur SEC HPLC then you have issues likely to be aggregate.

Most of what they are discussing is mostly inconsequential. If you get an IGF-1 test done on your serum and, in general if you get a rise from your baseline of 100 +/- 25 points points per iu then your GH is pretty close to well dosed. It varies some for people that have wacky metabolism, which are few or else the test would be useless. So if your baseline is 175 and you are on 2 iu a day you might see 375 and that should make you happy. Even if you see 300 you should be OK with that. If you test like Rocket does all the time and you use the same lab then you have a better grip on what your specific numbers are telling.

Purity is another issue. Anecdotally, side effects seem to be related to purity. Some US pharma grade GH seems to give less water retention side effects. That's something you are not going to be able to evaluate using a clinical serum test and probably will only rarely see a supplier post anything that will reveal anything interperable in that realm. You would need to see release specifications and clinical experience data to understand and interpret.

The guys in the pasted thread are amateurs trying to play professional drug regulatory reviewers in a black market with little to no data on products mostly made in China under god knows what kind of conditions.
__________________
See Glycomann's articles http://www.worldclassbodybuilding.com/forums/f497/
Reply With Quote
The Following 2 Users Say Thank You to Glycomann For This Useful Post:
blm (04-20-2017), chadd77 (04-20-2017)
Reply

Bookmarks


Currently Active Users Viewing This Thread: 1 (0 members and 1 guests)
 
Thread Tools Search this Thread
Search this Thread:

Advanced Search
Display Modes

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off

Forum Jump


All times are GMT -5. The time now is 09:09 PM.



Copyright ©2000 - 2018, Jelsoft Enterprises Ltd.
follow worldclassbodybuilding at facebook follow worldclassbodybuilding at pinterest follow worldclassbodybuilding at twitter