Ideality in Context: Motivations for Total Synthesis.

Total synthesis-the ultimate proving ground for the invention and field-testing of new methods, exploration of disruptive strategies, final structure confirmation, and empowerment of medicinal chemistry on natural products-is one of the oldest and most enduring subfields of organic chemistry. In the early days of this field, its sole emphasis focused on debunking the concept of vitalism, that living organisms could create forms of matter accessible only to them. Emphasis then turned to the use of synthesis to degrade and reconstitute natural products to establish structure and answer questions about biosynthesis. It then evolved to not only an intricate science but also a celebrated form of art. As the field progressed, a more orderly and logical approach emerged that served to standardize the process. These developments even opened up the possibility of computer-aided design using retrosynthetic analysis. Finally, the elevation of this field to even higher levels of sophistication showed that it was feasible to synthesize any natural product, regardless of complexity, in a laboratory. During this remarkable evolution, as has been reviewed elsewhere, many of the principles and methods of organic synthesis were refined and galvanized. In the modern era, students and practitioners are still magnetically attracted to this field due to the excitement of the journey, the exhilaration of creation, and the opportunity to invent solutions to challenges that still persist. Contemporary total synthesis is less concerned with demonstrating a proof of concept or a feasible approach but rather aims for increased efficiency, scalability, and even "ideality." In general, the molecules of Nature are created biosynthetically with levels of practicality that are still unimaginable using the tools of modern synthesis. Thus, as the community strives to do more with less (i.e., innovation), total synthesis is now focused on a pursuit for simplicity rather than a competition for maximal complexity. In doing so, the practitioner must devise outside-the-box strategies supplemented with forgotten or newly invented methods to reduce step count and increase the overall economy of the approach. The downstream applications of this pursuit not only empower students who often go on to apply these skills in the private sector but also lead to new discoveries that can impact numerous disciplines of societal importance. This account traces some select case studies from our laboratory over the past five years that vividly demonstrate our own motivation for dedicating so much effort to this classic field. In aiming for simplicity, we focus on the elusive goal of achieving ideality, a term that, when taken in the proper context, can serve as a guiding light to point the way to furthering progress in organic synthesis.

Preparation of Key Intermediates for the Syntheses of Coenzyme Q10 and Derivatives by Cross-Metathesis Reactions.

An alternative catalytic strategy for the preparation of benzylmethacrylate esters, key intermediates in the synthesis of coenzyme Q10 and derivatives, was reported. This strategy avoided undesirable stoichiometric reduction/oxidation processes by utilizing the catalytic formation of allylarenes and then cross-metathesis to selectively form E-benzylmethacrylate esters with good yields (58-64%) and complete E-selectivity. The ester intermediates were reduced to common key benzylallylic alcohols (90-92% yield), which were subsequently used in the formal syntheses of coenzyme Q10 and one derivative.

Development of hydrophilic gels containing coenzyme Q10-loaded liposomes: characterization, stability and rheology measurements.

OBJECTIVE: The aim of this study was to develop, characterize and evaluate stability of a gel containing coenzyme Q10 (Q10)-loaded liposomes, and enhance the stability of Q10 in the nanocarrier-containing gel compared to the conventional gel. METHODS: Q10-loaded liposome dispersions prepared from unsaturated or saturated lecithin, were characterized for particle size, polydispersity index (PDI), zeta-potential, pH value, oxidation index, Q10-content and morphology, and incorporated into carbomer gel. Liposome gels and liposome-free gel were analyzed for flow properties, pH values, Q10-content, and liposomes size and PDI (liposome gels), 48 h after preparation and in predetermined time intervals during 6 months storage at different temperatures in order to predict their long term stability. RESULTS: Liposomes were of small particle size, homogeneous, negatively charged, and their incorporation into gel did not significantly change (p > .05) their particle size and PDI. All gels revealed non-Newtonian, shear-thinning plastic flow behavior during storage with no marked changes in rheological parameters. Storage of gels did not significantly influence the pH value (p > .05), while it significantly decreased Q10-content (p < .05). Q10 was significantly more (p < .05) stable in liposome gel containing unsaturated lecithin liposomes (G1) than in gel containing saturated lecithin liposomes (G2) and liposome-free gel (G3). CONCLUSIONS: Q10-loaded liposome gel G1 was the optimal formulation, since during storage at different temperatures, it did not show significant increase in liposome size and PDI, it provided significantly higher stability for Q10 than other gels and its pH value was suitable for skin application. Due to limited Q10-stability it should be stored at 4 °C.

Characterization of Inclusion Complex of Coenzyme Q10 with the New Carrier CD-MOF-1 Prepared by Solvent Evaporation.

The aim of the current study was to evaluate the physicochemical properties of a solid dispersion of coenzyme Q10 (CoQ10)/cyclodextrin metal organic frameworks-1 (CD-MOF-1). As a result of the powder X-ray diffraction (PXRD), it was confirmed that the CD-MOF-1 was changed from the α form to the ß form by evaporation (EVP). A diffraction peak due to melting of CoQ10 disappeared the EVP (CoQ10/CD-MOF-1 = 1/2). The structure of this complex is presumed to be similar to the ß form of CD-MOF-1. As a result of the differential scanning calorimetry (DSC), the endothermic peak due to the melting of CoQ10 disappeared the EVP (CoQ10/CD-MOF-1 = 1/2). As a result of the near-infrared (NIR) absorption spectroscopy, findings suggested the hydrogen bond in formation between the CH group in the isoprene side chains of CoQ10 and the OH group of CD-MOF-1. Therefore, the formation of crystal solid dispersion in CoQ10/CD-MOF-1 was suggested. As a result of the dissolution test in distilled water, the EVP (CoQ10/CD-MOF-1 = 1/2) had better dissolution in comparison to CoQ10 alone. Furthermore, also in fasted state simulated intestinal fluid (FaSSIF) in vivo, the EVP (CoQ10/CD-MOF-1 = 1/2) had better dissolution in the human body than CoQ10 alone. From the results of 2D-nuclear overhauser effect spectroscopy (NOESY) NMR spectroscopy, CD-MOF-1 could not include the benzoquinone ring of CoQ10. It was confirmed that the isoprene side chain was included. Therefore, it was suggested that CD-MOF-1 useful as a novel drug carrier for CoQ10.

Synthetic Ubiquinones Specifically Bind to Mitochondrial Voltage-Dependent Anion Channel 1 (VDAC1) in Saccharomyces cerevisiae Mitochondria.

The role of the voltage-dependent anion channel (VDAC) as a metabolic gate of the mitochondrial outer membrane has been firmly established; however, its involvement in the regulation of mitochondrial permeability transition (PT) remains extremely controversial. Although some low-molecular-weight chemicals have been proposed to modulate the regulatory role of VDAC in the induction of PT, direct binding between these chemicals and VDAC has not yet been demonstrated. In the present study, we investigated whether the ubiquinone molecule directly binds to VDAC in Saccharomyces cerevisiae mitochondria through a photoaffinity labeling technique using two photoreactive ubiquinones (PUQ-1 and PUQ-2). The results of the labeling experiments demonstrated that PUQ-1 and PUQ-2 specifically bind to VDAC1 and that the labeled position is located in the C-terminal region Phe221-Lys234, connecting the 15th and 16th ß-strand sheets. Mutations introduced in this region (R224A, Y225A, D228A, and Y225A/D228A) hardly affected the binding affinity of PUQ-1. PUQ-1 and PUQ-2 both significantly suppressed the Ca2+-induced mitochondrial PT (monitored by mitochondrial swelling) at the one digit µM level. Thus, the results of the present study provided, for the first time to our knowledge, direct evidence indicating that the ubiquinone molecule specifically binds to VDAC1 through its quinone-head ring.

Meroterpenoids from a Medicinal Fungus Antrodia cinnamomea.

Antrodia cinnamomea, a medicinal fungus indigenous to Taiwan, has been shown to exhibit a broad spectrum of bioactivities for the treatments of alcoholic intoxication, diarrhea, abdominal pain, and fatigue, and a number of active principles have been identified. Among the bioactive entities, clinical trials of antroquinonol and 4-acetyl antroquinonol B are being carried out for curing cancer, hypercholesterolemia, and hyperlipidemia. The total synthesis of antroquinonol has been achieved; however, investigating the structure-activity relationship of this class of compounds remained difficult due to the lack of available analogues. Twenty antroquinonols isolated from A. cinnamomea IFS006 are reported herein. Their structures were elucidated using spectral analysis and by comparison with literature values. Of these, 11 antroquinonol analogues, namely, antroquinonols N-X (1-11), were previously unreported. The growth inhibitory activity of all the antroquinonol analogues was evaluated against human A549 and PC-3 cancer cell lines, and antroquinonol A exhibited the most potent activity, with GI50 values of 5.7 ± 0.2 and 13.5 ± 0.2 µM, respectively. Antroquinonols V (9) and W (10) also showed growth inhibitory activity against A549 cells with GI50 values of 8.2 ± 0.8 and 7.1 ± 2.1 µM, respectively, compared to 5-fluorouracil (GI50 = 4.2 ± 0.2 µM).

Reduction of Synthetic Ubiquinone QT Catalyzed by Bovine Mitochondrial Complex I Is Decoupled from Proton Translocation.

We previously succeeded in site-specific chemical modifications of the inner part of the quinone binding pocket of bovine mitochondrial complex I through ligand-directed tosylate (LDT) chemistry using specific inhibitors as high-affinity ligands for the enzyme [Masuya, T., et al. (2014) Biochemistry 53, 2304-2317, 7816-7823]. To investigate whether a short-chain ubiquinone, in place of these specific inhibitors, serves as a ligand for LDT chemistry, we herein synthesized a LDT reagent QT possessing ubiquinone scaffold and performed LDT chemistry with bovine heart submitochondrial particles (SMP). Detailed proteomic analyses revealed that QT properly guides the tosylate group into the quinone binding pocket and transfers a terminal alkyne to nucleophilic amino acids His150 and Asp160 in the 49 kDa subunit. This result clearly indicates that QT occupies the inner part of the quinone binding pocket. Nevertheless, we noted that QT is a unique electron acceptor from complex I distinct from typical short-chain ubiquinones such as ubiquinone-1 (Q1) for several reasons; for example, QT reduction in NADH-QT oxidoreduction was almost completely insensitive to quinone-site inhibitors (such as bullatacin and piericidin A), and this reaction did not produce a membrane potential. On the basis of detailed comparisons of the electron transfer features between QT and typical short-chain quinones, we conclude that QT may accept electrons from an N2 cluster at a position different from that of typical short-chain quinones because of its unique side-chain structure; accordingly, QT reduction is unable to induce putative structural changes inside the quinone binding pocket, which are critical for driving proton translocation. Thus, QT is the first ubiquinone analogue, to the best of our knowledge, the catalytic reduction of which is decoupled from proton translocation through the membrane domain. Implications for mechanistic studies on QT are also discussed.

Fluorogenic Ubiquinone Analogue for Monitoring Chemical and Biological Redox Processes.

We report herein the synthesis and characterization of a fluorogenic analogue of ubiquinone designed to reversibly report on redox reactions in biological systems. The analogue, H2B-Q, consists of the redox-active quinone segment found in ubiquinone, 2,3-dimethoxy-1,4-benzoquinone, coupled to a boron-dipyrromethene (BODIPY) fluorophore segment that both imparts lipophilicity in lieu of the isoprenyl tail of ubiquinone, and reports on redox changes at the quinone/quinol segment. Redox sensing is mediated by a photoinduced electron transfer intramolecular switch. In its reduced dihydroquinone form, H2B-QH2 is highly emissive in nonpolar media (quantum yields 55-66%), while once oxidized, the resulting quinone H2B-Q emission is suppressed. Cyclic voltammetry of H2B-Q shows two reversible, 1-electron reduction peaks at -1.05 V and -1.37 V (vs ferrocene) on par with those of ubiquinone. Chemical reduction of H2B-Q by NaBH4 resulted in >200 fold emission enhancement. H2B-QH2 is shown to react with peroxyl radicals, a form of reactive oxygen species (ROS) as well as to cooperatively interact with chromanol (the active segment of α-tocopherol). Kinetic analysis further shows the antioxidant reactivity of the nonfluorescent intermediate semiquinone. We anticipate that the H2B-Q/H2B-QH2 off/on reversible couple may serve as a tool to monitor chemical redox processes in real-time and in a noninvasive manner.

Efficient synthesis and antioxidant activities of N-heterocyclyl substituted Coenzyme Q analogues.

A new strategy for the efficient synthesis of C-5 heterocyclyl substituted Coenzyme Q analogues was developed by N-alkylation of bromomethylated quinone 11 with a series of amines 12 under metal-free conditions. In vitro antioxidant activities of these Coenzyme Q analogues were evaluated and compared with commercial antioxidant Coenzyme Q10 by employing DPPH assay. All these N-heterocyclyl substituted Coenzyme Q analogues are found to be exhibiting good antioxidant properties and may be used as potent antioxidants for combating oxidative stress.

Characterization of the reaction of decoupling ubiquinone with bovine mitochondrial respiratory complex I.

We previously produced the unique ubiquinone QT ("decoupling" quinone), the catalytic reduction of which in NADH-quinone oxidoreduction with bovine heart mitochondrial NADH-ubiquinone oxidoreductase (complex I) is completely decoupled from proton translocation across the membrane domain. This feature is markedly distinct from those of typical short-chain quinones such as ubiquinone-1. To further characterize the features of the QT reaction with complex I, we herein synthesized three QT analogs, QT2-QT4, and characterized their electron transfer reactions. We found that all aspects of electron transfer (e.g. electron-accepting activity and membrane potential formation) vary significantly among these analogs. The features of QT2 as decoupling quinone were slightly superior to those of original QT. Based on these results, we conclude that the bound positions of QTs within the quinone binding cavity susceptibly change depending on their side-chain structures, and the positions, in turn, govern the behavior of QTs as electron acceptors.

Synthesis of (+)-Antroquinonol: An Antihyperglycemic Agent.

The total synthesis of antroquinonol has been accomplished through Suzuki-Miyaura cross-coupling and Barton-McCombie reaction, and the α,ß-unsaturation was achieved through selenylation and oxidation protocols. In vitro and in vivo studies on the glucose-lowering properties of antroquinonol indicate that it is a potential antidiabetic agent.

A short synthesis of (±)-antroquinonol in an unusual scaffold of 4-hydroxy-2-cyclohexenone.

Antroquinonol, which was first isolated from a mushroom, Antrodia cinnamomea, found in Taiwan, is an anticancer compound with a unique core structure of 4-hydroxy-2,3-dimethoxycyclohex-2-enone carrying methyl, farnesyl and hydroxyl substituents in the 4,5-cis-5,6-trans configuration. A short synthesis of (±)-antroquinonol is accomplished in seven steps from 2,3,4-trimethoxyphenol, which is oxidized in methanol to a highly electron-rich substrate of 2,3,4,4-tetramethoxycyclohexadienone and then a Michael reaction with dimethylcuprate is performed as the key step, followed by alkylation, reduction and epimerization to incorporate the required substituents at three contiguous stereocenters.

Identification of the binding site of the quinone-head group in mitochondrial Coq10 by photoaffinity labeling.

Mitochondrial Coq10 is a ubiquinone (UQ)-binding protein that is a member of the steroidogenic acute regulatory protein (StAR)-related lipid transfer (START) domain superfamily. Deletion of the COQ10 gene was previously shown to cause a marked respiratory defect in Saccharomyces cerevisiae and Schizosaccharomyces pombe, which indicated that Coq10 may support efficient electron transfer between the respiratory complexes; however, its physiological role remains elusive. To elucidate the role of Coq10, we attempted to identify the binding site of UQ in recombinant S. pombe Coq10 expressed in an Escherichia coli cell membrane through photoaffinity labeling with the photoreactive UQ probe, UQ-1, in combination with biotinylation of the labeled peptide by means of the so-called click chemistry. Comprehensive proteomic analyses revealed that the quinone-head ring of UQ-1 specifically binds to the N-terminal region of Phe39­Lys45 of Coq10, which corresponds to the ligand-binding pocket of many proteins containing the START domain. The labeling was completely suppressed in the presence of an excess amount of artificial short-chain UQ analogues, such as UQ2. In the Phe39Ala and Pro41Ala mutants, the extents of labeling were ∼40 and ∼60%, respectively, of that of wild-type Coq10. While Coq10 has been thought to bind UQ, our work first provides the direct evidence of Coq10 accommodating the quinone-head ring of UQ in its START domain. On the basis of these results, the physiological role of Coq10 has been discussed.

Total synthesis of (±)-antroquinonol d.

Total synthesis of (±)-antroquinonol D, which is isolated from very expensive and rarely found Antrodia camphorata and which has potential anticancer properties, was achieved from 4-methoxyphenol. In addition, a Michael addition to dimethoxy cyclohexadienones was studied. The main step involved chelation and substrate-controlled diastereoselective reduction of cyclohexenone and lactonization. Lactone synthesis facilitated the diastereoselective reduction of ketone, which help control the desired stereochemistry at the crucial stereogenic center in the natural product. Other key reactions in the synthesis involved a Michael addition of dimethyl malonate on cyclohexadienone, dihydroxylation, and Wittig olefination. A sesquiterpene side chain was synthesized through coupling with geranyl phenyl sulfide and Bouveault-Blanc reduction.

Effects of cytoprotective antioxidants on lymphocytes from representative mitochondrial neurodegenerative diseases.

Two new aza analogues of the neuroprotective agent idebenone have been synthesized and characterized. Their antioxidant activity, and ability to augment ATP levels have been evaluated in several different cell lines having suboptimal mitochondrial function. Both compounds were found to be good ROS scavengers, and to protect the cells from oxidative stress induced by glutathione depletion. The compounds were more effective than idebenone in neurodegenerative disease cells. These novel pyrimidinol derivatives were also shown to augment ATP levels in coenzyme Q(10)-deficient human lymphocytes. The more lipophilic side chains attached to the pyrimidinol redox core in these compounds resulted in less inhibition of the electron transport chain and improved antioxidant activity.

Synthesis and characterization of mitoQ and idebenone analogues as mediators of oxygen consumption in mitochondria.

Analogues of mitoQ and idebenone were synthesized to define the structural elements that support oxygen consumption in the mitochondrial respiratory chain. Eight analogues were prepared and fully characterized, then evaluated for their ability to support oxygen consumption in the mitochondrial respiratory chain. While oxygen consumption was strongly inhibited by mitoQ analogues 2-4 in a chain length-dependent manner, modification of idebenone by replacement of the quinone methoxy groups by methyl groups (analogues 6-8) reduced, but did not eliminate, oxygen consumption. Idebenone analogues 6-8 also displayed significant cytoprotective properties toward cultured mammalian cells in which glutathione had been depleted by treatment with diethyl maleate.

Efficient Lewis acid ionic liquid-catalyzed synthesis of the key intermediate of coenzyme Q10 under microwave irradiation.

An efficient synthesis of a valuable intermediate of coenzyme Q(10) by microwave-assisted Lewis acidic ionic liquid (IL)-catalyzed Friedel-Crafts alkylation is reported. The acidity of six [Etpy]BF(4)-based ionic liquids was characterized by means of the FT-IR technique using acetonitrile as a molecular probe. The catalytic activities of these ionic liquids were correlated with their Lewis acidity. With increasing Lewis acid strength of the ionic liquids, their catalytic activity in the Friedel-Crafts reaction increased, except for [Etpy]BF(4)-AlCl(3). The effects of the reaction system, the molar fraction of Lewis acid in the Lewis acid ILs and heating techniques were also investigated. Among the six Lewis acid ionic liquids tested [Etpy]BF(4)-ZnCl(2) showed the best catalytic activity, with a yield of 89% after a very short reaction time (150 seconds). This procedure has the advantages of higher efficiency, better reusability of ILs, energy conservation and eco-friendliness. The method has practical value for preparation of CoQ(10) on an industrial scale.

Assembly and Characterization of Biocompatible Coenzyme Q10 -Enriched Lipid Nanoparticles.

Coenzyme Q10 (CoQ10 ) is a strongly hydrophobic lipid that functions in the electron transport chain and as an antioxidant. CoQ10 was conferred with aqueous solubility by incorporation into nanoparticles containing phosphatidylcholine (PtdCho) and apolipoprotein (apo) A-I. These particles, termed CoQ10 nanodisks (ND), contain 1.0 mg CoQ10 /5 mg PtdCho/2 mg apoA-I (97% CoQ10 solubilization efficiency). UV/Vis absorbance spectroscopy of CoQ10 ND revealed a characteristic absorbance peak centered at 275 nm. Incorporation of CoQ10 into ND resulted in quenching of apoA-I tryptophan fluorescence emission. Gel filtration chromatography of CoQ10 ND gave rise to a single major absorbance peak and HPLC of material extracted from this peak confirmed the presence of CoQ10 . Incubation of cultured cells with CoQ10 ND, but not empty ND, resulted in a significant increase in the CoQ10 content of mitochondria as well as enhanced oxidative phosphorylation, as observed by a ~24% increase in maximal oxygen consumption rate. Collectively, a facile method to solubilize significant quantities of CoQ10 in lipid nanoparticles has been developed. The availability of CoQ10 ND provides a novel means to investigate biochemical aspects of CoQ10 uptake by cells and/or administer it to subjects deficient in this key lipid as a result of inborn errors of metabolism, statin therapy, or otherwise.

Enhancing regiocontrol in carboaluminations of terminal alkynes. Application to the one-pot synthesis of coenzyme Q10.

Two new "generations" of methodological advances are reported for the Negishi carboalumination of terminal alkynes. Use of simple, inexpensive additives that alter the Al-Zr complex formed between Me(3)Al and Cp(2)ZrCl(2) give rise to an especially effective reagent mix that results in virtually complete control of regiochemistry upon carboalumination of 1-alkynes. One timely application to coenzyme Q10 is highlighted. Regioisomers from subsequent coupling, which would otherwise be very difficult to separate, are avoided.

Radical-scavenging polyphenols: new strategies for their synthesis.

New strategies for the synthesis of polyphenols, compounds with antioxidant properties contained in every kind of plants, are discussed. Syntheses of different classes of polyphenols, namely ubiquinones, present in many natural systems in which electron-transfer mechanisms are involved, hydroxytyrosol, one of the main components of the phenol fraction in olives, and flavonoids, widespread in the plant kingdom, were approached by simple and environmentally sustainable methods.