Saturday, May 16, 2015

Reader Question: Steroidal Glycosides in Solanum Maternum Fruit

The tamarillo fruit is the fruit of a subtropical Solanum shrub
A reader contacted me via email recently posing a very intriguing question regarding the levels of steroidal glycoalkaloids present in a close relative of the tamarillo (tree tomato). I've reproduced an excerpt of his correspondence below and I ask any readers with practical experience in this area or with academic interest in characterizing the steroidal natural products in this wild fruit to get in touch with me ( so that I can relay the information to the relevant parties.
Hello Brian,

For context, I grow a wild South American Solanum species, Solanum maternum, which I doubt has ever been investigated for the glycoalkaloids present (it was discovered relatively recently).

It is the (extremely) close relative of the 'Tamarillo' ('tree tomato'), Solanum betaceum (synonym Cyphomandra betacea), a fruit in limited international commerce, and also grown here in New Zealand.

While in potatoes the 'normal' level of glycoalkaloids of 20-100 mg/kg is said to be 'not of concern' - albeit a 'safe' level has not been established - introducing genes from wild species of potato may inadvertently introduce undesirable higher levels of glycoalkaloids. In addition, new glycoalkaloids, 'untested' in human diet, may be introduced.

Solanum maternum has total resistance to the fungus disease powdery mildew; S. betaceum, the fruit of commerce, is susceptible. S. maternum is of interest for breeding for disease resistance - just as are wild potato species for the domesticate.

Fruit of S. maternum have a mix of acidity, unusual floral flavor notes - and a savage bitterness and throat-gripping 'acridity' that render it inedible. The Tamarillo has significant bitterness in the fruit wall, but not in the pulp cavity, and it is the pulp only that is eaten. Nevertheless, past consumer testing has shown some people report a 'catching' in the throat when they eat Tamarillo.

I haven't been able to find any internet information on the glycoalkaloid load in tamarillos, but assume, based on their historical wide acceptance, that it is relatively low. I assume that S. maternum has a 'high' glycoalkaloid load. There may be other chemicals present in the maternum fruit that adds to the general 'acridity' or 'heat'. But it has never been investigated...Checking the internet (I have no background in chemistry), I see there are some simple reagents which give a color precipitate in the presence of glycoalkaloids. An industrial chemist might make one or more for me, but these do not seem to be able to give a quantitative indication of levels, which is really what I am after. This is a faint hope - but I have to ask - do you know of any newly invented 'field kit' for a rough quantitative indication of glycoalkaloid levels? Alternatively, do you think there would be sufficient academic interest in characterizing the steroidal complexes in this wild fruit (ideally, from my point of view, in comparison with the domesticate, and also in comparison with fruit from a cross of the two species)?

Regiodivergent Outcomes in Metal-Catalyzed Aliphatic C-H Azidations of Estrone Derivatives

            New late-stage C-H functionalization reactions that are compatible with structurally complex and clinically useful natural products are of great interest to the pharmaceutical industry. This is due, in part, to the ability of nitrogen-containing functionality to modulate the binding affinity and physicochemical properties of drug candidate molecules. However, until recently, chemical methods to convert an alkyl (and, in particular, a tertiary) C-H bond into an alkyl carbon-nitrogen bond were lacking. This problem is further exacerbated by the fact that enzymes generally do not catalyze aminations of C-H bonds (for an exception to this rule, see this recent report). Earlier in 2015, two new synthetic methods for the metal-catalyzed azidation of aliphatic C-H bonds were received with great interest and enthusiasm by the chemical community. Interestingly, although the C-H azidation methodologies are mechanistically related, their application to differentially protected estrone derivatives produces regiodivergent outcomes (see Scheme above). In one case, the benzylic tertiary position (C9) reacts primarily and, in the other, the secondary C6 C-H position is functionalized (for details, see below).
            The method reported by the laboratory of John Groves at Princeton University is a manganese-catalyzed process that uses sodium azide as the azide source. Mechanistically, this chemistry is analogous to Groves’ previously disclosed manganese-catalyzed C-H fluorination wherein a fluoride ligand axial to manganese transfers fluorine to a carbon-centered radical derived from the hydrocarbon substrate. For azidation, the fluoride is simply replaced by an azide source (NaN3). The substrate radical is then trapped by the in situ-generated Mn(IV)-azide complex to construct the carbon-nitrogen bond. Both manganese porphyrins as well as Mn salen-type Jacobsen catalysts are competent participants in the catalytic cycle and the novel azidation protocol can be run under air. Under the optimized reaction conditions, estrone acetate is converted predominantly to a C9-a-azide. A diazidation product wherein both benzylic positions (C6 and C9) are functionalized is also observed as a major side product.
            John Hartwig’s group has also developed a method for late-stage azidation of tertiary and benzylic C-H bonds using an iron catalyst and Zhdankin’s hypervalent azidoiodinane reagent. For a nice introductory overview of Hartwig’s technology, see this blog post. Azidation of TBS-protected estrone, under the iron-catalyzed conditions, furnishes the corresponding C6-a-azide in modest yield. The stereodivergent nature of the two related azide-forming processes is somewhat striking.
            On a somewhat unrelated note, the laboratory of Timothy Newhouse at Yale University has disclosed a new palladium-catalyzed methodology for a,b-dehydrogenation of esters and nitriles. The method nicely complements (and, in some cases, exceeds) earlier, more classical approaches to achieve a,b-unsaturation such as the Saegusa-Ito oxidation or the Sharpless selenoxide elimination (later extended by Grieco). The novel reaction from the Yale group is compatible with nitrile substrates derived from estrone (shown above), cholesterol and androstenedione-type steroids.

Tuesday, April 21, 2015

Baran’s Semisynthetic Work on Ouabagenin and Related Bioactive Glucocorticoid Derivatives

            Phil Baran’s laboratory at The Scripps Research Institute recently disclosed a full account of their 20-step synthesis of the aglycone of ouabain starting from a derivative of cortisone. Their general strategy for introduction of the dense hydroxylation pattern of the target molecule was to ‘redox-relay’ pre-existing oxidation states that are naturally present within the cortisone framework to neighboring and distal carbon atoms. For example, using Norrish type II photochemistry, the C11 ketone oxidation state was effectively relayed to the angular C19 methyl group through the intermediacy of a cyclobutanol. Oxidative fragmenation of the cyclobutanol then furnishes the C19-functionalized product. Baran’s optimization and extension of the Norrish protocol to the problem of functionalization of the unactivated steroid C19 substituent provides a useful and complementary alternative to more classical tactics, such as those reported years ago by Barton, as well as Meystre and Heusler from Ciba. A more detailed analysis of Baran’s partial synthesis of ouabagenin can be found here.

            Interestingly, Baran’s group describes a foray into the realm of medicinal chemistry in their recent JACS full article. They report the application of the Norrish type II photochemical C-H functionalization methodology to the synthesis of C19-hydroxylated analogues of corticosteroid drugs such as clobetasol propionate. The authors indicate a motivation to minimize the undesired biological effects of glucocorticoid receptor (GR) agonists stemming from off-target mineralocorticoid antagonism. Unfortunately, the novel clobetasol analogue 1, synthesized in eleven steps from cortisone acetate, exhibits binding affinity to GR that is inferior to clobetasol propionate by about an order of magnitude. While selectivity against the mineralocorticoid receptor is not reported, 1 does exhibit moderate anti-inflammatory efficacy in a cellular assay. Baran’s analogues are clearly lacking 9a-halogenation. The 9a-halogenated glucocorticoid series was first described in 1953 and has been shown to provide a >10-fold increase in binding affinity to GR, relative to parent C9-proteo hormones. The synthetic route used to prepare clobetasol analogue 1 is shown below.
            Arguably, the most robust and practical methodology reported by Baran for semisynthetic cardenolide modification with applications to therapeutically relevant corticosteroids was disclosed much earlier. In the Journal of Organic Chemistry, Baran reports a strategy for systematic deoxygenation of ouabain as a means to obtain important 17b-hydroxyacetyl pregnane-type steroids. Ouabain, by virtue of its pre-built oxygenation at the C11 position and latent side chain derivable from the C17 butenolide, is a logical candidate for a synthetic precursor to various glucocorticoids.
            As proof of this principle, ouabagenin 1,19-acetonide was reductively deoxygenated (see below) at the C1 position via the intermediacy of a 3,11-dione. Subsequently, upon exposure of an 11,19-methyl-ketal to thionyl chloride and pyridine at low temperature, the tertiary hydroxyl group at C14 eliminates to generate an olefinic intermediate. Catalytic hydrogenation of this intermediate results in a saturated system with b-hydrogen atoms located at the newly formed stereogenic ring junction positions. Finally, oxidative manipulation of the a,b-unsaturated lactone moiety affords a 20-oxo-14-iso-pregnane that can be considered a 19-hydroxylated congener of cortisone acetate. Most of the synthetic operations described in this manuscript were demonstrated on decagram scale. For example, the hydrogenation depicted below (step 3) was conducted on 73-gram scale and, ultimately, 4 grams of the 14-isopregnane glucocorticoid derivative were generated from a single batch run.
            This latter synthetic work was conducted by John S. Baran (i.e. not Phil) of The Laboratories of G. D. Searle and Co., Chicago, Illinois. The single author manuscript (see excerpt below) was received by JOC on August, 29, 1963. During this time, the post-World War II era race to synthesize cortisone was in full swing. Ouabain and strophanthidin were two cardenolides that were evaluated as feedstock precursors to the newly discovered and highly sought-after corticosteroid wonder drugs of the 1950s – 1960s. G. D. Searle, much like other major players in the cortisone race - Merck, Syntex and Upjohn - figured prominantly in the vitally important steroid research that was ongoing during this time period.