Periodic Reporting for period 4 - BioMet (Selective Functionalization of Saturated Hydrocarbons)
Berichtszeitraum: 2023-05-01 bis 2024-10-31
Selective functionalization of unreactive C–H bonds represents a significant paradigm shift from traditional organic synthesis, which typically focuses on reactive functional groups. Extending undirected C–H bond functionalization necessitates new methodologies capable of selectively activating similar C–H bonds in substrates and products without cross-reactivity. Historically, selective functionalization of alkanes without directing groups has been particularly challenging, especially for undirected activation of primary C–H bonds. Interest in this field is driven by the potential to sustainably valorize inexpensive alkanes, primary constituents of natural gas and petroleum. Although transition metal-based transformations have shown promise in selective alkane functionalization, they often remain limited to specific functionalization types. A general, high-yielding approach for selectively activating primary C–H bonds and enabling diverse terminal functionalizations remains elusive. Our ERC research developed a strategy transforming linear alkanes into alkenes, followed by remote functionalization using a metal-catalyzed hydrometalation/migration sequence. This sequence generates primary organometallic intermediates, which react with electrophiles to produce exclusively primary-functionalized alkanes.
Hydrocarbon activation via dehydrogenation is challenging, typically requiring high temperatures and leading to side reactions. We explored biocatalytic desaturase systems, as enzymes offer high selectivity under mild conditions. A mutant bacterial strain, developed through random mutagenesis, hyperproduced internal alkenes from linear alkanes. Optimizing the reaction conditions using phosphate buffer, monosodium glutamate, thiamine•HCl, and magnesium sulfate enabled the conversion of hexadecane into cis-hexadecene regioisomers with a 61% isolated yield and 83% dehydrogenation efficiency.
This fermentation reaction was scaled for multigram synthesis and applied to C14–C20 alkanes, producing exclusively (Z)-isomers. The microorganism also facilitated the desaturation of aliphatic xenobiotics and terminally functionalized derivatives. Substrates such as n-hexadecyl methyl ether and halogenated alkanes yielded α-functionalized Z-olefins, demonstrating high chemoselectivity and functional group tolerance.
With reliable mono-unsaturated hydrocarbon derivatives in hand, we focused on remote, selective terminal functionalization. This strategy employs transition metal complexes for long-range chain migration, yielding anti-Markovnikov terminal organometallic intermediates. Hexadecenes were subjected to various metal-mediated hydrofunctionalization reactions, achieving excellent site-selectivity (α > 98:2). Terminal alkylboranes were synthesized using pinacolborane and a cobalt pincer complex. Reactions with trichlorosilane and chloroplatinic acid yielded trichloroalkylsilanes, which were converted into trimethyl- and trivinyl-hexadecylsilanes.
Hydrozirconation, another valuable approach, enabled broad electrophilic functionalization. Treating biogenerated alkenes with Cp2ZrCl2 and Red-Al produced terminal hexadecylzirconocene chloride intermediates, which were electrophilically trapped to yield 1-halogenohexadecanes. Oxidation produced cetyl alcohol, while reactions with hydroxylamine derivatives yielded primary amines. Reactions with TMSCN provided n-heptadecanonitrile in 75% yield. Copper(I)-catalyzed reactions enabled C–C bond formation, and nickel-catalyzed systems achieved exclusive sp3–sp3 cross-coupling.
This unified two-step strategy yielded a broad range of functionalized derivatives, including halides, silanes, boranes, alcohols, amines, nitriles, and C–C bonded products. Following these achievements, we initiated a study to elucidate the bacterial dehydrogenation mechanism. RNAseq analysis identified candidate genes, validated by qPCR, genetic engineering, and whole-genome sequencing. Comparative genomic analysis against related strains lacking dehydrogenation capability and outer membrane protein isolation further refined our understanding.
Future efforts include genetic engineering to enhance conversion efficiency and substrate scope by overexpressing key pathways, increasing gene copy number, and engineering more efficient enzymes.
We also investigated the metal-walk process to understand its transformation mechanism. A modular approach was developed to synthesize acyclic products with distant stereocenters in various relationship via a metal-walk, completed in three catalytic steps from commercially available alkynes. This approach enabled efficient diastereocontrol and high E:Zratios, crucial for natural product synthesis, as demonstrated in the formal synthesis of the α-tocopherol side chain.
This fermentation reaction was scaled for multigram synthesis and applied to C14–C20 alkanes, producing exclusively (Z)-isomers. The microorganism also facilitated the desaturation of aliphatic xenobiotics and terminally functionalized derivatives. Substrates such as n-hexadecyl methyl ether and halogenated alkanes yielded α-functionalized Z-olefins, demonstrating high chemoselectivity and functional group tolerance.
With reliable mono-unsaturated hydrocarbon derivatives in hand, we focused on remote, selective terminal functionalization. This strategy employs transition metal complexes for long-range chain migration, yielding anti-Markovnikov terminal organometallic intermediates. Hexadecenes were subjected to various metal-mediated hydrofunctionalization reactions, achieving excellent site-selectivity (α > 98:2). Terminal alkylboranes were synthesized using pinacolborane and a cobalt pincer complex. Reactions with trichlorosilane and chloroplatinic acid yielded trichloroalkylsilanes, which were converted into trimethyl- and trivinyl-hexadecylsilanes.
Hydrozirconation, another valuable approach, enabled broad electrophilic functionalization. Treating biogenerated alkenes with Cp2ZrCl2 and Red-Al produced terminal hexadecylzirconocene chloride intermediates, which were electrophilically trapped to yield 1-halogenohexadecanes. Oxidation produced cetyl alcohol, while reactions with hydroxylamine derivatives yielded primary amines. Reactions with TMSCN provided n-heptadecanonitrile in 75% yield. Copper(I)-catalyzed reactions enabled C–C bond formation, and nickel-catalyzed systems achieved exclusive sp3–sp3 cross-coupling.
This unified two-step strategy yielded a broad range of functionalized derivatives, including halides, silanes, boranes, alcohols, amines, nitriles, and C–C bonded products. Following these achievements, we initiated a study to elucidate the bacterial dehydrogenation mechanism. RNAseq analysis identified candidate genes, validated by qPCR, genetic engineering, and whole-genome sequencing. Comparative genomic analysis against related strains lacking dehydrogenation capability and outer membrane protein isolation further refined our understanding.
Future efforts include genetic engineering to enhance conversion efficiency and substrate scope by overexpressing key pathways, increasing gene copy number, and engineering more efficient enzymes.
We also investigated the metal-walk process to understand its transformation mechanism. A modular approach was developed to synthesize acyclic products with distant stereocenters in various relationship via a metal-walk, completed in three catalytic steps from commercially available alkynes. This approach enabled efficient diastereocontrol and high E:Zratios, crucial for natural product synthesis, as demonstrated in the formal synthesis of the α-tocopherol side chain.
The objective of this work was to establish a general principle in organic synthesis for generating a broad range of complex molecular architectures from simple saturated hydrocarbons. Our strategy involves an initial transformation of the alkane into an alkene, which then undergoes a metal-catalyzed hydrometalation/migration sequence. The first step—essentially the removal of two hydrogen atoms—is accomplished using a genetically engineered bacterial strain. Importantly, the position and geometry of the resulting double bond do not influence the second step, as the metal-catalyzed hydrometalation/migration efficiently isomerizes the double bond along the carbon chain to selectively yield the primary organometallic species. Subsequent trapping of these organometallic intermediates with a variety of electrophiles produces the desired functionalized alkane.
This work led to the development of novel, selective, and efficient processes for utilizing simple hydrocarbons, unlocking the synthetic potential of raw hydrocarbon feedstock for the environmentally sustainable production of new compounds and materials. Additionally, an in-depth biological mechanistic study was carried out to enhance the efficiency of the bacterial strain, ultimately overcoming some existing limitations.
In parallel, we conducted extensive research on the metal-walk transformation to gain a deeper understanding of its constraints and systematically address them. This transformation has now evolved into a broadly applicable method in organic chemistry, enabling selective functionalization at distant positions along a carbon chain with complete stereochemical control.
This work led to the development of novel, selective, and efficient processes for utilizing simple hydrocarbons, unlocking the synthetic potential of raw hydrocarbon feedstock for the environmentally sustainable production of new compounds and materials. Additionally, an in-depth biological mechanistic study was carried out to enhance the efficiency of the bacterial strain, ultimately overcoming some existing limitations.
In parallel, we conducted extensive research on the metal-walk transformation to gain a deeper understanding of its constraints and systematically address them. This transformation has now evolved into a broadly applicable method in organic chemistry, enabling selective functionalization at distant positions along a carbon chain with complete stereochemical control.