Hydroxylation at a chosen carbon

Given an ordinary organic molecule and one carbon–hydrogen bond marked in advance, a single written procedure replaces that hydrogen with a hydroxyl group and does not do the same at the other carbon–hydrogen bonds. The marked carbon may be one the usual catalysts would rather leave alone.

A hydroxyl group is how a lead compound becomes a metabolite, a handle for a conjugate, or the oxidation state a natural-product route spends several steps reaching. Those steps disappear only if the site can be nominated. A method that oxidizes whichever C–H bond is already the most reactive does not change that practice.

Two small-molecule catalysts set the present standard, and both select the site themselves.

Gormisky and White reported non-heme iron catalysts that enhance or overturn a substrate’s innate preference for aliphatic C–H oxidation, in preparative yields, together with a model that ties the outcome to electronic, steric, and stereoelectronic parameters of the sites (doi:10.1021/ja407388y (opens in a new tab)). The supporting information of that paper states that primary sites are uniformly not oxidized under those conditions and are excluded from the model. The predicted site is the one the model ranks, not a carbon marked by an outside laboratory.

Chan, Palone, Bietti, and Costas later hydroxylated the primary C–H bonds of tert-butyl groups with 1 mol percent of a manganese catalyst and hydrogen peroxide in nonafluoro-tert-butyl alcohol (doi:10.1002/anie.202402858 (opens in a new tab)). On 2,2,3,3-tetramethylbutane the primary alcohol formed in 51 percent yield by gas chromatography, beside 2 percent of the aldehyde, for 96 percent selectivity of the alcohol over further oxidation. Replacing acetic acid with 4 equivalents of urea raised the combined yield of oxidation products to 71 percent, with 96 percent product selectivity and a mass balance above 99 percent. Two further hydrocarbons gave the primary alcohols in 60 percent and 53 percent yield by the same assay, at or above 91 percent product selectivity, and only the primary site reacted even though secondary C–H bonds were present. A capecitabine-derived carbamate gave the primary alcohol in 28 percent isolated yield, with 39 percent of the starting material recovered. The same paper shows the preference slipping as the tert-butyl group moves away from an electron-withdrawing group: the ratio of tert-butyl oxidation to oxidation at other sites is 1.3 for the C6 pivalate. These are catalyst-selected sites on substrates chosen for the study.

No report opened for this problem takes an arbitrary marked carbon on a blinded set, writes the procedure before any experiment on those molecules, and returns the alcohol at that carbon.

An independent laboratory preregisters 20 molecules. Each has a molecular weight between 150 and 500, at least four constitutionally distinct carbon–hydrogen sites, and no protecting group that is not already part of the structure as drawn. On each structure the laboratory marks exactly one carbon. The marked carbon bears at least one hydrogen. The required product is the alcohol at that carbon, not the ketone, the aldehyde, or the carboxylic acid.

At least six of the twenty marked carbons are disfavored, by a rule fixed before the structures are released. A marked carbon is disfavored only when all three of the following are true: it is a methyl or a methylene; it is not benzylic, not allylic, and not adjacent to oxygen, nitrogen, or sulfur; and the same molecule contains at least one other carbon–hydrogen site that is either a tertiary aliphatic C–H or a benzylic C–H. The laboratory records which six meet the rule. The attempt sees the marked structures and can apply the same rule.

The structures are revealed together. Within 24 hours, and with no experiment on any of the 20 molecules, the attempt submits one written procedure. Only the substrate changes from molecule to molecule. The laboratory then runs each procedure once, as written, on 0.2 to 0.5 mmol of substrate. A second procedure may be submitted in the same 24 hours and is used only if the first fails to produce isolable material. No third procedure is allowed, and nothing is changed after the first spectrum.

The isolated material is characterized by high-resolution mass spectrometry, a proton spectrum, a carbon spectrum, and an HSQC and HMBC pair sufficient to assign the site of hydroxylation. The regioisomer ratio is the proton-NMR integral of the specified alcohol divided by the sum of the integrals of every other monohydroxylated isomer in the crude mixture, measured before chromatography against a weighed internal calibrant.

The attempt passes when at least 16 of the 20 substrates give the specified alcohol in an isolated yield of at least 30 percent, with a regioisomer ratio of at least 10 to 1, and at least 4 of those 16 are among the 6 disfavored sites.

The count, the 30 percent yield, the 10-to-1 ratio, and the 4-of-6 disfavored requirement are pass marks defined by this problem. They are not claimed as records. Thirty percent is below the 51 percent gas-chromatography yield of the primary alcohol, and below the 71 percent combined oxidation yield, that Chan and co-workers report on a hydrocarbon whose reactive site is the one their catalyst prefers. It is there so the product is isolated and assigned rather than detected. Ten to one means the specified site accounts for at least about 91 percent of the monohydroxylated material. The disfavored-site count is what puts the bar past a method that only oxidizes the tertiary or benzylic C–H bond the substrate already offers.

A no-reagent blank, identical except for the absence of the catalyst or reagent named in the procedure, is run for two of the twenty substrates, chosen by the laboratory before the procedures are opened. One of the two is disfavored. The blank must not produce the specified alcohol above 2 percent yield by the same NMR assay. If it does, the batch is invalid and is repeated with a cleaned setup.

Site assignment is done by a chemist who did not write the procedure, using only the spectra and the starting structure. If that chemist cannot place the hydroxyl group, the substrate counts as a failure. Over-oxidation of the specified alcohol to a ketone or carboxylic acid does not count as the alcohol.

Any reagent, catalyst, enzyme, light source, or electrochemical cell that the written procedure names. A directing group that is already present in the preregistered drawing may be left in place. Computation before the 24-hour clock starts is allowed. A different catalyst for a disfavored site is allowed only if that choice is written in the procedure submitted before any experiment on the set.

Installing a directing group, a halogen, a boron substituent, or any other functional handle on the way to the alcohol. A separate protection step that is not in the preregistered structure. Screening conditions on the test substrate, or on a close analog made for the purpose, after the structures are revealed. Counting a ketone or a hydroperoxide as the alcohol. Using a different substrate that already bears the hydroxyl group. Treating a disfavored site as optional and reporting only the fourteen ordinary sites. A different published method for each marked site. One written procedure applies to every substrate, with only the substrate changed. The single backup, if it is used, applies to every substrate or to none.

At least 10 of the 20 substrates give the specified alcohol in at least 15 percent isolated yield, with a regioisomer ratio of at least 5 to 1, at least 2 of the 10 are disfavored sites, and the blanks pass. Partial success is not a solution. Fifteen percent, 5 to 1, and 2 of 6 are pass marks for this lesser bar only.

The 16-of-20 count, the 4-of-6 disfavored requirement, the 30 percent yield, the 10-to-1 ratio, the 2 percent blank ceiling, and the partial-success figures are pass marks defined here.

Cited measurements:

• Gormisky and White, Journal of the American Chemical Society, 2013, 135, 14052–14055, doi:10.1021/ja407388y (opens in a new tab). Catalyst-controlled aliphatic C–H oxidation in preparative yields, with a model of innate site preference. The supporting information states that primary sites are not oxidized under those conditions and are excluded from the model.
• Chan, Palone, Bietti, and Costas, Angewandte Chemie International Edition, 2024, 63, e202402858, doi:10.1002/anie.202402858 (opens in a new tab). The 51 percent, 2 percent, 60 percent, 53 percent, 71 percent, 96 percent, 91 percent, 28 percent isolated, 39 percent recovered, and 1.3 site ratio quoted above are taken from that paper.

Purity, where a purity number is reported alongside the isolated yield, is measured by the absolute quantitative proton NMR assay of Pauli and co-workers, Journal of Medicinal Chemistry, 2014, 57, 9220–9231, doi:10.1021/jm500734a (opens in a new tab).