basic field tests for alkaloid content in leaf, stem and root tissue

[Frut]

Hello

Looking for a simple field test or procedure to determine (approximately) total alkaloid content in a range of wild plants in situ. Starting with Bala-sida sp.

Must be light weight and simple and not cost prohibitive. Hopefully relatively rapid as well.

Procedure wanted to obtain a ball park estimate of total alkaloid content in the field, so that individual genotypes can be compared and more interesting individuals located again if desired. Cant control soil effects on total alkaloid content, that is understood, but will note soil type on soil surface at each sample specimen, as this data could yield a useful relationship to alkaloid content. By keeping the procedure as rapid and simple as possible, hopefully can minimise variability due to time between sampling individuals, ie do a lot of plants in one area in one hit.

Maybe can get a rough handle on 24 hr fluctuation in a few individuals as well and extrapolate to larger sample to minimise this type of noise when comparing the whole data set (ie previous point raised on temperature changes in the one day can make a big difference in the same individual plants alkaloid content).

Cant see any legal issues, as not wanting to do or obtain anything illegal, just get a handle on the abundant natural resources in my region, of a legal plant and a legal family of alkaloids, Australians had to do this in a previous crisis, when cut off in the second world war and they done this well. Cant see any law that prevents us making ourselves just as ready today, except if the plant is illegal. Come what may economic crisis, environmental crisis or military crisis or ones own health crisis it pays to know the resources on your doorstep, does it not?

best regards

Pedro Frut

[Chiral]

This is a huge task to undertake. With the plant being such a rampant weed, one would think hypothetically that it would be best to mass harvest many plants at a time, then process them all as a single batch and collect whatever alks are available.

To wander around and do alkaloid testing in the bush on thousands of plants and tag them, well I just don't see the need as there is so much of it growing, there will be slight variations across many plants but not significant to worry about.

The less time one has to spend walking around gathering and testing these plants the better.

[Frut]

Thanks Chiral

This is a huge task to undertake.

Depends how large I want to make the sample size and sampling area

With the plant being such a rampant weed, one would think hypothetically that it would be best to mass harvest many plants at a time, then process them all as a single batch and collect whatever alks are available.

Yes, if my goal was to collect alkaloids ASAP, but my goal at this time is only to get a handle on the amount of variability between different individuals of the same species and see if there are any obvious patterns such as

alkaloid content and

ie relationship to soil type, time of day, season, etc

To wander around and do alkaloid testing in the bush on thousands of plants and tag them, well I just don't see the need as there is so much of it growing,

I dont know about testing thousands, hundreds maybe of the more vigorous specimens, but you can see why my query is princiapally to find a simple field test, rapid and giving a rough idea in situ. I would only tag and gps more outstanding individuals. How simple was that UV light that use to be for sale on SAB? Did one have to pick stuff and take it home to do at night etc? or could it be done in daylight with the UV light? perhaps there is something better?

I am not looking to do rocket science in the field, just isolate a few more promising individuals if they exist, so I can come back later an focus just on a few plants.

there will be slight variations across many plants but not significant to worry about.

Is this an assumption Chiral? on what basis do you suggest this?

The less time one has to spend walking around gathering and testing these plants the better.

I dont know, I dont mind wandering around my local bush looking at things. Please dont assume I want to get out there and extract things ASAP, so I can have a lot of this or that extract or alkaloid at hand ASAP. It is not my goal.

Currently I am more interested in observing things. I certainly would like to hone down and road test a procedure here that may be applicable to quite a range of species around the globe, and I hope may be of interest and practical application to other ethnobotanists and medicinal / psychotropic plant enthusiasts. Naturally I want to do this honing down in a way that causes no harm and yields the maximum benefit and appreciate all input to this end.

regards

Pedro Frut

Edited December 2, 2009 by Frut

[Chiral]

The variations across plants are going to occur with plants due to mass, exposure, water, age, general health etc etc...there are no two plants identical in the wild and this is what I'm referring too really. You could perhaps find out by drying and weighing, you will need an equal amount of plant material roughly say from one patch and then another, but to try and find out if one plant say 6' away from another at roughly the same size has more alkaloid than the other is well...I'm not Ethno field guide or expert but I would assume the differences are going to quite neglible. testing the seed, stem, flowers, leaves and roots is one way but sheesh fair amount of work...I admire you enthusiasm that's for sure.

When you have finished doing this and completed your studies, would you mind doing some DNA testing/studies, alk profiling on all the Trichocereus cactus in Australia and put us all out of our misery...

[Alice]

Good stuff Frut, you've set yourself quite a goal. Sounds like a good project, I wish I had that sort of time on my hands.

And I'd just like to say, I think it's excellent that you're starting this off with a weed. I can only imagine the environmental impact that large scale sample collecting might have on rare/endangered species.

[Alice]

Oh, and as far as field tests go, for something portable and relatively quick/easy, a basic tlc kit and some spray reagents for spot tests (Marquis maybe, or similar) would be the way to go. Though I'm not sure that you would get the kind of specificity between similar alkaloids that you might be after from a spot test (although the Rf of the spot, combined with the colour, would get you very close).

Thats about all you can do as far as field tests go, even the fed's portable HPLC/GCMS is hardly an instrument you can fit in a backpack.

Edited December 2, 2009 by Alice

[MORG]

I'm not a chemist so not much use to you in regard to the practical here.

What you intend to do is very interesting though. I am becoming more and more interested in plant secondary metabolites and I have found through recent reading that there is a strong bias in the literature towards a very narrow sample because of a focus on biosynthesis and molecular biology.

A fundamental step in studying the evolutionary and ecological role of alkaloids I think is to study them at a population level. This is so rarely found in the literature. Characterizing variation within and between populations are very worthy observations to make.

Good luck!

And I should add: Studying weeds makes perfect sense in terms of ethics and logistics!

Edited December 2, 2009 by MORG

[ENtiTY]

Affraid I'm a bit skeptical as to the usefulness of such an undertaking on the basis of Chiral's post.

Momomoto: I believe the sida in your pic to be Sida spinosa. I have both rhombifolia and spinosa in my back yard and have taken a pic to show the differences between the 2.

The piece on the left is spinosa; has more defined serations on the leaves, is a paler green, lacks the velvety whitish texture on the undersides of the leaves and tends to have a low to prostrate habit.

In the middle and on the right are rhombifolia; part of the leaf has no serration, is a darker green, leaves tend to be elongated although they can be a rounded shape as well, underside of leaf has a whitish velvety texture and tends to grow upright in a small shrub habit.

I included 2 pieces of rhombifolia to demonstrate the variation in leaf shape.

The right hand leaf on the spinosa branchlet is actually face down so you can see the underside of the leaf and observe the lack of velvety whiteness. Similarly the right hand middle leaf on the center rhombifolia branchlet is showing the underside of the leaf.

There are more differences but the ones I mentioned above are obvious and simple to use for identification.

Sorry to hijack the thread but I believe a pic of a sida that was posted by Momomoto in the previous version of this thread that is now locked was mis-identified.

[t st tantra]

any spines on the spinosa?

t s t .

[planthelper]

thx, harry!

that's a fantastic pic and explanation, awesome post!

having read your reply, i will not miss id them again, hehe.

i like the idea aswell and it would be very interressting to know if fluctuations of alkaloid % do excist in wild populations.

some chemist might know if some field alkaloid test can produce a result telling us which plant contained more activas.

[Alice]

some chemist might know if some field alkaloid test can produce a result telling us which plant contained more activas.

Doubtful. You would need to collect large amounts of material, extract, ID, weigh and calculate yields for the active. Not a field job.

You'd need a lot of material so that you get a decent, weighable amount of alkaloid. This is especially important when you're looking to compare plants that might only have between 0.05-0.1% alkaloid DRY weight. Unless the plant is quite big, you'd probably need to harvest the whole thing to determine an accurate % alkaloid.

However, you may be able to get a rough idea qualitatively by looking weighing the leaves/whatever to say 5g, do a quick field extraction using say 5 mL of solvent, spotting onto your tlc plate and using a visualising agent to show the spots. As long as you keep the masses of material, volume of solvent, and number of spots the same, you should be able to get a rough idea by the darkness of the spot or the rapidity of development (think pill testers, mmm instant black!). You may need to concentrate the extract before spotting though. It wont give you a % but should give a ball park idea of super-strains.

[Alchemica]

Here's a few field test reagents (excludes a few interesting alkaloids but it's a start): http://www.ncjrs.gov/pdffiles1/nij/183258.pdf

OK for detecting the presence of some of the alkaloids but I'm not sure how easily you could distinguish the level of actives merely via colour intensity/development time when spotting on a TLC plate.

You should be able to precipitate most alkaloids from extracted solutions as certain salts/complexes (tannates, picrates, phosphomolybdates, platinum complexes, potassium mercuric iodide complexes etc) and use the precipitates to get a total alkaloid percentage. Still, not overly field friendly or easy.

[reptyle]

whats the average dosage of sida rhombifiolia?

[Frut]

Thanks all, for your ideas and insights

I will attempt to discuss some aspects of these replies as my mind tries to digest them

The old adage of there being many ways to skin a cat comes to mind but perhaps saying many ways to boil a yage brew would be better as I have a respect for cats, mixed with a fear too.

I think when my head starts spinning when I get this type of feedback I have to go back to my original question, and the context in front of me, then consider your suggestions.

My Question(s) as it currently stands after your comments:

1. Are there individual Bala-sida genotypes in my immediate region that have a significantly higher total alkaloid content

2. What factors are driving any observed variability in alkaloid content, are they genetically driven or environmentally driven (ie soil, nitrogen content) or what is the relative impact of genetics and environment in this alkloid content.

3. Are there predictable patterns of variability in the individual over time in the same tissues, or between different tissue compartments like roots versus stems at a given time.

I think tackling questions on the alkaloid makeup (actives inactives etc) is largely relevant to question 2 and 3 type levels of enquiry, and something to be done mostly back in the lab with larger harvested samples, this comes to mind when I read comments from Alchemia, Chiral and the suggestions of using drug test kits. Holy catfish this is potentially a great left field idea to look into Alchemia, which is partly why I am posting about this on this board, as I suspect there is a quite a few of us here that are left field (to use the jargon) that is when we are on the field at all.

As Chiral points out time is the essence, I guess too Chiral I will be drawn to those Bala individuals that I perceive to be a good agricultural form. More biomass in a nice manageable shape. This is inevitable. It could be a scrawny little 6 leaf runt and have an alkaloid content from hell or heaven? but it will most likely go missed. But lets face it, plants that punch out secondary compounds that are a high metabolic cost on that plant, make this effort usually because they have some vigour left over in the kitty after the food, clothes and shelter is paid for. if you are struggling to find a feed of beans each day you are not likely to find time and resources to buy a powerful new weapon to keep the bugs and thugs away. So going for those individuals that visually stand out in ones immediate perception from a group is not such a stupid thing, perhaps? Its the normal brain filter and perhaps a dam useful one for once?? Look! there is a fucken big green one over there with a lot of flowers!

If I have to sample living tissue in situ with a gram or so in a test tube, some solvent and a reagnet well whats the bloody simplest thing guys. I really dont need to isolate, anthropromorphic active and inactive alkaloids, at this stage do I?

I just want to answer this at first! is this a plant that may be worth coming back to for a second look? ie to take a bit bigger sample for something, like take back to lab? Something like alkaloids present yes.....a little, usual amount, or yes more! gps that dude! move on

Does this thinking make sense?

What is this marquis and TIC and spot stuff Alice?, sorry for my ignorance here, does it give you a ball park estimate of total alkaloid in situ and not weigh half a ton or turn me into a walking solvent bomb! Somebody mentioned chloroform, that fucker doesnt burn easy does it? but I dont want to enter a state of mind where I sample 1 ala pronto perhaps sample 2 takes the rest of ther day, I guess just do it down wind.

I dont have time for mountainous studies either folks and this is why I am picking your brains for short cuts. Have you not on your travels, kept striking some plant of intense intrest that the locals are cultivarting or its coming up as a weed everywhere and wanted to get a handle on which were the better indiviudals from your position? do they exist? what would be the easy way perhaps funnest way to have half a chance on coming up with something real here?

Chiral, Trichocereus genus or whatever they call it now surely is good one to work on, the DNA stuff too though it sounds even less field easy and the cost.....?

When I said the format we are developing here has potential for wider application, I did not envisage destructive harvesting of any rare and endangered species to run field or lab alkaloid tests etc. To me that is insane, and I hope it is to others, I imagine it would be on this board. Shit if a plant is that scarce whats the point in putting an effort into the kinds of questions I am suggesting here. The question in that context surely must be how do we protect this unique treasure (even if we humans have no economic use for it!) how do we give it a chance to reestablish itself into a viable population in the wild in its natural habitat (more than often then it follows that its whole habitat has gone anyway and it gets more and more depressing).

I am in a place with large wild resource of the invasive bala-sida, S cordifolia I think (but Harry now has me wondering so I better up some photos soon for your ID) By the way Harry the more you hijack this thread with posts of your quality the better please feel free for that was a real useful wake up call on ID and my basic assumptions on the species and genus. There seems enough evidence to suspect this plant as a whole extract, or part extract ie root or stem tissue may contain a whole complexity of medicinally useful compounds for humans and some veterinary applications. Other evidence points to this Bala-sida plant genus potentially containing phytochemical compounds that may have applications or uses beyond medicine to industrial and agricultural uses including bio-mining. This plant may not be a waste product after all? From a hard edged agricultural economist perspective it largely depends on just how many usefuls are in the plant overall and how easy/cheap are they to be comverted into useful endproducts?

I dont want to be a hard edge agr. economist but I do want to keep my feet somewhat near ground level and somewhere near the field be it left side or right or whatever. Though I will steer clear of the dudes in the centre of the field if possible they might be agricultural economists or regulators and after all I want to actually do something that may be a half useful pilot study even if the sample size is too small it may tantalise for bigger things?

thanks people

Pedro Frut

ps I wish I had a good sense of smell or taste, a simple taste and yes that one is bitter level 5 or 4 or 1 would be sufficient! but it wouldnt be a very widely applicable technique for alkaloid studies unless they were shortish in a short career.

[Alice]

What is this marquis and TIC and spot stuff Alice?, sorry for my ignorance here, does it give you a ball park estimate of total alkaloid in situ and not weigh half a ton or turn me into a walking solvent bomb! Somebody mentioned chloroform, that fucker doesnt burn easy does it? but I dont want to enter a state of mind where I sample 1 ala pronto perhaps sample 2 takes the rest of ther day, I guess just do it down wind.

I dont have time for mountainous studies either folks and this is why I am picking your brains for short cuts.

 

Hmm, seems that you're not too familiar with organic chemistry, I would suggest jumping on wiki and looking up

TLC

retention factor

marquis reagent

This is the very least you need to know to even begin your project.

There is no quick and easy quantitative field test that will achieve what you want. It's going to take a lot of work.

[MORG]

You're looking at a potential PhD worth of research right there. Ambitious and I encourage you.

If you're serious about disentangling the genetic and environment influences on alkaloid production you should consider a common garden experiment. It is really the only effective way to look at this. If you want to go deep, breeding experiments and common garden will allow you to calculate heritability coefficients (the extent to which a trait is inherited versus environmentally controlled). There are textbooks written on quantitative genetics, all about determining the genetic/environmental interaction on phenotype.

I don't know much about growth rate/generation time in Sida to know if this is at all feasible for you.

[planthelper]

this is a very big project, and i think frut is a bit unaware how difficult this type of field and lab work realy would be.

i would keep it very simple at the beginning and if results are interressting i would go back and do more work with a better (more precise) setup.

one thing not mentioned is that, you would have to make all samples, the same, as in regards from where from the plant those samples have been taken and ho developed the tissue was.

in other words one would have to take great care to take allways say same size of developed leaves from say close to the top, or something similar. i mean don't mix fan leaves with growing tips or so on.

i guess you could, forget the alkaloid field test, because we do know that those plants contain them, no point reestablishing that fact. we only came down this pathway because you thought this test will tell you aswell about the ammounts of alkaloids, but as it's now established it's not.

alchemicas idea with the precipitates is i thin a very good one, and the only feasable one.

let's assume frut is very skillfull but has little extraction knowledge, and has to decide between a full acid base extraction or a field solvent precipitation, i guess the latter is far less work.

i would like to know how this precipitation methode is done.

my guess is though that all sida plants will have similar alkaloid levels, and enviromental factors are the main source of variation of the activas.

another thing is that, because this tests are done by hand, you would get different results just because of your inconsitancy. so it would be paramount to stick to a manual.

[apothecary]

First up, Welcome to the forum Frut.

Doubt you have cordifolia judging by your location, much more likely to be rhombifolia. Considering the origins of cordifolia, the distribution of the plant in QLD makes more sense than to find it where you are. I haven't spotted a single cordifolia specimen south of NNSW, and even around there is heavily infested with rhombifolia.

I didn't want to add much because this thread really borders on the dodgy line to me, but why hasn't anyone mentioned steam distillation as the quickest method for quantification?

Just run your field with the roadside ephedrine/pseudoephedrine test to mark individuals of interest (because not every specimen will have what you want - I promise) and then return in season to harvest some root bark for distillation.

Some questions you need to ask yourself:

* Do I have the correct identification knowledge of this plant

* Do I have the chemistry know-how to attain my goal in a scientifically rigorous manner

* Am I liable to get into a lot of trouble undertaking this study without the aegis of an educational institution (the answer is almost certainly yes)

Edited December 7, 2009 by apothecary

[reptyle]

This looks like the best party i've ever seen!

 

as for dodgy...IT'S SCIENCE~~~~~

Obviously you know something about the scientific method frut...more controls than i would have thought to even contemplate...

if anyone can read this...i'm pretty sure that frut isn't looking for your condonation or any sort of comment about how big and daunting this research might be, as that was already identified by frut in one of fruts posts...

as far as the question goes...WHAT IS A RELIABLE WAY OF TESTING ALKALOID LEVELS, NAMELY EPHEDRINE AND PSEUDO-EPHEDRINE IN THE FIELD...are there any ethnobotanists out there that might have an idea???

there's nothing wrong with legitimate biological research, especially when it has the goals and possible results have already been identified and are quite legitimate...

grow up SAB...eeek. It seems like everybody hates their reflection so much they attempt to "save" each other through misguided advice regarding "risks" and possible "trouble" that might arise as a result...errr!

alchemists made this world...paranoia is for sick people. and fear is not a natural response!

are there any pH type reagents that are specific for certain types of alkaloids?

How do the Gov't test for speed, compared to cocaine compared to heroin compared to baking powder and how then do they test for purity or concentration to ascertain whether it is product or simply contamination...it's not a long and drawn out process...

peace.

Edited December 7, 2009 by reptyle

[reptyle]

i take it all back...i love you guys...the war is over.

i think i'm slightly frustrated...i tried to make a box, for one of those drums out of the other thread...and the edges don't line up...i suck at using a circular saw.

going to do some glueing and nailing...

Edited December 7, 2009 by reptyle

[reptyle]

sorry for the distraction from the real science...

[Alchemica]

Just a way that you could run your tests with minimal investment using simple, home equipment and a free program utilising "Background subtracted image density" (BSID) measurements (can send you the pdf if desired)

 

 

 

Quantitative and Qualitative TLC Analysis (edited for this forum):

 

 

 

We are reporting here our method for qualitative and quantitative analysis of the titled compounds in addition to a unique photometric method for quantitative analysis of UV and visualized chromaphores.

 

 

 

Using TLC plates we have determined the rF values of the titled compounds and identified their wet and dry chromaphores using the modified Van Urk/Salkowski reagent system as well as their respective concentrations.

 

 

 

Methods:

 

 

 

Silica Gel 60 F254 hard surface(polymer) 2.5x7.5 cm plates were used as the TLC sorbent system.

 

 

 

Reagent System:

 

 

 

The reagents were made a follows.

 

 

 

(A) van Urk37 reagent: 1 g p-dimethylaminobenzaldehyde was dissolved in 50 ml conc. HCl (specific gravity 1.190) and 50 ml absolute ethanol was added;

 

 

 

( B ) Salkowski6 reagent 2.03 g FeCI3. 6 H2O were dissolved in 500 ml water and 300 ml conc. H2SO4 (specific gravity. 1.840); this reagent is stable indefinitely.

 

 

Spray reagent. The new TLC spray reagent used, was made up of reagent A and B (1 :3).

 

 

 

We also tested the method previously published of using a cotton ball to swipe the plate rather than spraying. The method that was found most efficacious was to hold the saturated cotton ball in one hand and the top of the plate(hard polymer, gypsum will notwork with this method) and swipe the plate vigorously in one stroke. Some gray artifacts may appear on the plate but these disappear upon drying and do not affect the analysis. This method avoids the need for a sprayer and fume hood.

 

 

 

A single plate was spotted with Alkaloid 2x and the chloroform/methanol/water (84:14:1) solvent system was tested as was the Propanol/Water/Amm Hydroxide (8:1:1). As you can see from the following photograph under short-wave u.v. the Propanol system provided an expanded spot size and concentric rings while the chloroform spot stayed relatively intact. Based upon this analysis we selected the Propanol system.

 

 

 

2 samples of alkaloid were available. 25 mg of each sample was diluted in 5 ml of Methanol. A 5 λ pipette was used, which gave 25 μgrams of material per spot. Spotting had to be done repeatedly to avoid spot from growing too large. In our new photographic method, it becomes very important to mark the exact center of the loading spot at the start point of the plate, as measurements are in pixels and often times it is difficult to get the spot right on the cross hairs that are drawn with pencil.

 

 

 

Based upon the solvent front length average measurement of ____ mm and the front spot edge average of ____ mm we get an average rF from the two measurements of XXX

 

 

 

Using the Background subtracted integrated density(BSID) measurement in the image tool, we find that the absolute value for the 10 lambda spot equals _____. This is the average of two measurements.

 

 

 

The average abs value for the 5 lambda spot = _____. Dividing the 10 lambda measurement by the 5 lambda measurement we get a value of ______

 

 

 

Thus by using two or more measurements of the area (these are drawn by hand so multiple measurements are suggested) we get a result of _____. Thus we have proven that the Image tool can accurately measure the relative concentration of the alkaloid on the TLC plate and therefore is an accurate tool for qualitative measurement of TLC plate concentrations!

 

 

 

Now we have, with a simple digital camera, Short-wave U.V. lamps and fluorescent TLC media, the equivelant of $20-30k lab quality analytical instruments. TheUTHSCSA Image tool can be downloaded for free from the site

 

 

http://ddsdx.uthscsa...g/download.html.

 

 

 

Attempts to analyze the wet image, show below were not successful as the wet plate does not have the fully developed density of the dry plate and the measurements were in error.

 

 

 

Note the typical dry colors of the alkaloid as _____ with a dark circle around the spot. This is the standard color for this compound.

 

 

 

Average BSID measurements of these two spots gave the following results:

 

 

 

Find Average 10 lambda BSID measurement

 

 

 

Find Average 5 lambda BSID measurement

 

 

Find Ratio of 10 lamdba/5 lambda spots

 

 

 

Thus we have demonstrated the accuracy of the Image tool in analyzing dry plates when plates have been visualized using the Van Urk/Salkowski reagent system.

 

 

Conclusions:

 

 

Qualitative measurements of TLC plates are now possible using simple, home equipment. Utilizing a free program, Image, Background subtracted image density(BSID) measurements, when averaged, correlate very well with the loadings on the plate. Use of a wet plate will not provide accurate measurements but will provide some qualitative information as to the nature of the materials under analysis. A 5 microliter spot containing 25 micrograms of the title compound was compared to a 10 microliter spot containing 50 micrograms of the title compound. The results for both U.V and visualized BSID measurements were ____ and ____ respectively.

 

 

 

Short Wave UV measurements of the image as well as dry visualized measurements provide accurate information as to the total amount of compound present when compared to a standard. This method now provides the experimenter with a rapid, easy way to quantify the amounts of compounds present in a sample and may be correlated to other compounds of interest to provide rapid, accurate measurements of your materials. This system compares favorably to analysis done using $20k –30k systems.

 

 

 

 

------

 

 

 

 

For the precipitation method:

 

 

It's worth getting the book Some micro-chemical tests for alkaloids (Stephenson) (around as an online ebook), too. You'd want to find the precipitate that was least soluble to get the best idea of the amount of alkaloid. Something like a sintered glass gooch filter of known weight and a vacuum filtration setup would be a handy way to collect/weigh the precipitates.

 

 

 

 

Otherwise, if anyone here at SAB wants to pay $4,000, you can buy portable analysis units the size of a shoe box from Scientrific that use air as a carrier and a data logging system to give retention times and peak areas... tempted to get one myself! Not sure how good they are.

 

 

Edited December 20, 2009 by Alchemica

[sascacheuan]

A good vid of "how to" Tlc:

 

[Frut]

Hello folks

I am sorry for not getting back to you for a while

First of all thank you once again for all your generous gift of thoughts and ideas and experiences relating to my driving question

The main factors driving alkaloid production in Sida-Bala

Whether your comments are perceived as negative or positive, condoning such an experiment or not, I do appreciate your feedback and I am both surprised and grateful for the wealth of knowledge expressed by so many people on this board

As to my apology of slowness I have been snowed under up to Xmas at the mundane end of keeping bread on the table, so had to put it on hold.

First I must get a handle on my plant ID (thanks Apothecary and Harry) Sida cordifolia or rhombifolia or?.

I am starting to get a feeling that there may be a bit of a Eucalypt phenomena where the actual individual Sida species that have ranges that overlap and similar morphologies may have somne interbreeding going on so we have fuzzy overlaps between species.

This is another question and typical of what appears to begin as simple and practical enquiries into natural systems. Ask one question end up in a Fibinacci or fractal type sequence of thousands.

I hear you, the fact I an opening a Phd worth of work. Sweet Lord I hope not! But that’s why I consult you dudes and fellow plant people that like to keep their fingers in the dirt and the journey practical. I don’t like tapping at computers and staring at them for hours on end and this Sida thing is a hobby interest to me.

Remember, I just want this to be a pilot look into the question of what is driving the alkaloid variability. I want it to be kept on the practical for we gardeners. Such as 1-4

1. Can we increase alkaloid or other pertinent phyto-chemical loads with simple soil ameliorations, site selection, timing and is it worth buggering around with genetic strain selection etc.

2. What can I find on this from the smallest amount of field samples and related tests and the smallest number of samples in my garden trial?

If the answer at the end is ‘hey yes’ first soil/microclimatic it may be worth beefing up nitrogen, growing on clay versus sand, or adding a clay component to sand etc, rhombifolia has the edge over cordifolia at my home site,

or even the contrary..viz… needs far more data and far more work to extract a pattern from the noise, then I would be satisfied for now

2: temporal: Harvest at flowering, harvest at seed set, or harvest before flowering etc. yields more of X or X or X

3: plant compartmental: main stems or roots or leave and smaller stems, take this one and use the rest as mulch etc. yield more of X or X or X

4: genetic, use rhombifolia or use cordifolia don’t fuck with the rhombis etc

Sure to do it really precisely accurately and sufficiently to show reliable repeatable patterns it involves and absolute shit load of data, probably 100 phds not 1 or 0.001 phd’s. But I subscribe to the philosophy of if it is worth doing at all? It is worth doing poorly at first!

My context

Wild resources

I have easy access to three large wild samples, firstly on sandy soil around Newcastle coastal regions, secondly on heavy clays up the Hunter Valley and Sydney’s west-Cumberland plains

Thirdly Hunter river silts that use to be sites that wild hemp was eradicated from in the 60-70’s, quite thoroughly it seems. Quite rich soils so largest Sida bushes on this silt as expected.

Cultivation resources

I have a little patch of earth that has been fed up on compost for more than 20 years, it is sandy (high silica, water repellent despite all the compost), slightly alkaline and I have lots of Sida growing just for their capacity to mine the subsoil and bring lost nutrients back up to the top.

My action plan for now, thanks to you guys

1. Species identification: ASAP

You are probably right Apothecary, it being rhombofolia and not cordifolia, Harry first lifted my suspicions here and as a result my camera is in getting repairs so I can get a few decent pics from the sample in the thread ASAP and get your feedback as they are in flower at full swing as I write (please watch this space my ident gurus Harry and Apothecary and others out there)

2. Gather in some wild samples now they are flowering, make repeatable with gps. Put these on hold by drying these or would it be better to freeze them fresh? Or better still to freeze briefly then juice through a Champion extractor then re-Freeze and save the labelled juice for later analyses?

My original question to you on how to do even the most basic simple test in the field to determine a maybe yes or no: This bush is likely to contain more of something that looks like alkaloids and this does not ?

Surely guys there must be something simple I can do? That doesn’t involve me going to the cops and asking where I get one of there breath or road swab kits?!!! At this level I don’t care what fucken combo of alkaloids it is, or even definitely yes this is alkaloids versus glycosides etc. Just a positive or negative at this stage on the question alkaloids possibly present or not, a maybe is sufficient???

Off course being able to get a handle at this point on whether it is a little bit of something that may be alkaloids are present, or a relative lot of alkaloids that may be alkaloids are present would be great.

Just so I know which samples I can take back to the lab (it seems a University may be coming on side to provide better lab facilities too for the later, I will let you know)

Reptyle said get the stuff, like the cops are using to determine (at your car window?) if this driving dude has been on smack, krank, X or gunja in the last n-hours. Alright I was critical above but it is a clever thought, for we suspect there may be phenylethylamines in the Sida, but the fact that they are none of these illicits, and these tests may be more sophisticated than necessary to pick it up, for they determine people on X from quite specific types of krank, I gather they don’t rake in the legit users of legal krunks like those conventionally treated for ADHD and narcolepsy, or the cops will be getting into a increasingly busy period of false positivos I imagine. My initial take on this, Reptyle, is it is too dam sophisticated for the purpose. And it also touches on issues with Sida where I really don’t want to go on this thread, to the cop shop asking where I can get this tool kit. It is not my interest with this plant, not my purpose with the plant, and dam unfortunate too for the study (more on this later) that it possibly contains traces of Xzephri- drunes.zX in many cases.

Guys how about the old UV light, why is Torsten not selling these lights any more? Surely what could be simpler than eyeballing a beam on a flattened sample of leaf, or stem or root in a dark room. Sure it is not on the spot, sample site, daylight time stuff, but this is where the cheap modern gps could make it easy to keep repeatable and lets say 50 specimens may be done in a few hours in the evening after a mornings work wandering along the Hunter River bank etc?

Please dudes? Why are we not talking UV, bouncer/nightclub door checker flashlights, or something really basic like that? Any of you guys out there used the ones that Torsten use to sell for a purpose like this?

Also thanks to Alchemia and Sascacheuan for such detailed info and the link re TLC, I am trying to see how I can use this easily in the field, or at home without driving to a uni lab, but I am dam sure it is useful back in the lab

Now for some important Housekeeping

Moderator(s) should we move this thread to chemistry or another less accessible part of the Corroborree site, as I don’t want to attract attention and interference from the krank spanners or people chasing krank spanners (krank spanner = people used by krank, not people that use krankas a tool to get a desired outcome).

I must admit I was a bit naive at first when posting my ideas on this. I did not realise the implications of discussing any plant that contains even the smallest trace of alkaloids in the XEphizdrineXvvvv family.

After trying to get my head around why my first thread was closed and people whom I have respect for on this site quickly gave me warnings mentioning words like jail, I first said on reading WTF?. I was shocked.

I personally was drawn to this plant after I found it had amazingly deep roots like Lucerne, and was more tough than Lucerne. Lucerne has revolutionized my garden, using it in green manure cycles, I can show you the before and afters. I am now mulching with Sida and cycling it through beds and it seems to be doing the same with a twist. I think it is drawing more trace elements, probably less nitrogen than Lucerne to the surface. My citrus leaves have quickly lost all the lacking zinc iron and mn look for example.

Also at heart I am still the apprentice alchemist, like many here, always looking for the gold bio-miner too!

Also a heavy mulch of Sida seems to drive my grape and fig tree cuttings into a more vigorous tap root structure from the word go! Which I am sure some of you would understand the significance of, on sandy soils. So I tried with last years fig and grape cuttings in pots (I sell quite a few)- putting a 60 mg XXXZpsuedo-z-ephedrinze-XXXzz pill on 15 pots (one tab per pot-broken up at weekly intervals) versus Sida mulch and compost tea on another 15 versus 15 samples in control (osmocote but same soil mix). Results:

Sida mulch and compost tea: very vigourous downward root growth,

Control, usual growth, moderate vigour, entirely adventitious growth.

OTC 60 mg generic XXXsidafudXz pills X 1 weekly I got something in between the above.

I personally doubt that any serious lawn enforcementi agency, dealing with the drouge problem would prioritise this Sida issue. I am told by a source in big pharma, whom (you may not agree, but I say give them their due) are very clever with chem type Locks, that big underbanana operations now days import their key feeds to what I imagine is a tricky and costly reduction process. I cant ever see Sida competing, I may be wrong, but the time and expense I imagine would always make the importation in this country the option when only profit makes it attractive to these types. I may be wrong.

But even if I am right I want to keep this thread entirely out of this sphere as much as possible, not due to the law or serious villains, as much as the fwack-nackles and also genuinely sad cases, that I feel a measure of compassion for, that want to put rocks in their heads or have already done so.

I thought this was happening due to the mods wisely closing the first thread and giving the chance for me to get off on a better foot, but was surprised to get a few pms recently (which I will ignore) from people I imagine from the krunk spanner class, whom I reckon could hinder progress towards some useful preliminary output.

So folks, and mods what do you suggest here (and a thanks to those that have forced me to get up to scratch on this issue, surrounding any poor plant that has even the smallest trace of Zxephri-dongeXz , in it, please keep reminding me if I slip from time to time).

I really think many of us need a simple processes and appropriate equipment to do pilot field studies on a range of plants for the exact same questions I am, proposing here and trying to keep the thread on. Unfortunately I have chosen a plant that may have a very small quantity of zXephi-drox-Zx family alkaloids in it, amongst a hell of a lot of other perhaps more useful synergistic stuff that to me is equally if not far more exciting, how do we keep this thread functional and not dragged down by krunk elements I now recognise that I was being warned I may attract?

Thanks all

Pedro Frut

Ps Incidentally doing some background reading on the above last issue, makes me wonder about the Xzephid-rinxXz molecule, how does a little plant tie so much energy into such a molecule? I base this on why that molecule needs literally, as I now understand, a bombs worth of energy to cleave off the single little (is it hydroxyl) side chain, to make some kind of krunk. Subject any other organic molecule to this, even a diamond you would blow it to smithereens in seconds surely? but this guy can take hours of blasting, almost like fission fuel requirements. Surely that is an interesting side issue? That meek little weeds make molecules harder than diamonds

Are all alkaloid molecules put together with such potent bonds? My god the energy requirements for a plant to do this? And why? there must be a pay off to that plant?

Just a little side thought, off topic, but thanks dudes for getting me to read up here as it makes me even more curious what sida and all the other disparate metabolisers of ZXephi-drubeXz have in common to warrant such a huge metabolic cost to make something harder than a diamond, pest disease protection, downward tap root growth stimulation? There must be some-things we are missing here Ummm back to my question:

The simplest way to say yay! Alkaoids may be present!, or not and hopefully

There may be more in this one less in that one…gps it please Pedro wantabee Boffin…..full stop!

Edited December 27, 2009 by Frut

[Darklight]

I hear you, the fact I an opening a Phd worth of work. Sweet Lord I hope not!

Mwahahahaha- you so are.

Guys how about the old UV light

There was a wealth of info by a bloke named Fractal here about alkaloid field tests using the UV light and TLC. The search engine can't seem to find them, maybe they went missing in an earlier upgrade? If anyone has copies of the posts or can find them on the SE, put the link here. The info in question was detailed but the process facile once you had all the gear

cheers

Darklight

Ah- here's some of it- keep looking for more

http://www.shaman-australis.com/forum/index.php?showtopic=2621&st=0&p=21647&fromsearch=1entry21647

Edited December 27, 2009 by Darklight