13‐Oxo‐9(Z),11(E),15(Z)‐octadecatrienoic Acid Activates Peroxisome Proliferator‐Activated Receptor γ in Adipocytes

Peroxisome proliferator‐activated receptor (PPAR)γ is expressed in adipose tissue and plays a key role in the regulation of adipogenesis. PPARγ activators are known to have potent antihyperglycemic activity and are used to treat insulin resistance associated with diabetes. Therefore, many natural and synthetic agonists of PPARγ are used in the treatment of glucose disorders. In the present study, we found that 13‐oxo‐9(Z),11(E),15(Z)‐octadecatrienoic acid (13‐oxo‐OTA), a linolenic acid derivative, is present in the extract of tomato (Solanum lycopersicum), Mandarin orange (Citrus reticulata), and bitter gourd (Momordica charantia). We also found that 13‐oxo‐OTA activated PPARγ and induced the mRNA expression of PPARγ target genes in adipocytes, thereby promoting differentiation. Furthermore, 13‐oxo‐OTA induced secretion of adiponectin and stimulated glucose uptake in adipocytes. To our knowledge, this is the first study to report that 13‐oxo‐OTA induces adipogenesis through PPARγ activation and to present 13‐oxo‐OTA as a valuable food‐derived compound that may be applied in the management of glucose metabolism disorders.     

13‐Hydroxy‐9Z,11E‐Octadecadienoic Acid Production by Recombinant Cells Expressing Burkholderia thailandensis 13‐Lipoxygenase

Linoleate 13‐lipoxygenase from Burkholderia thailandensis was expressed in Escherichia coli for the production of 13‐hydroxyoctadecadienoic acid (13‐HODE), an antiseptic emulsifier. Linoleate 13‐lipoxygenase in cells had higher thermal stability than the purified enzyme. To increase 13‐HODE production, recombinant cells were permeabilized by solvents, detergents, salts, and other chemicals. The enzymatic activity in cells was the highest for permeabilized cells treated with 0.5 M NaCl among the permeabilizers tested. The optimal reaction conditions for the production of 13‐HODE from linoleic acid by permeabilized cells treated with 0.5 M NaCl were at pH 7.5, 25 °C, 20 g/l linoleic acid, 15 g/l cells, 0.15 mM Cu²⁺, and 6 % (v/v) methanol in a 100‐ml baffled flask containing a 5‐ml working volume with agitation at 200 rpm. Under these conditions, permeabilized cells produced 15.8 g/l 13‐HODE after 30 min with a conversion yield of 79 % (w/w) and a productivity of 31.6 g/l/h. The conversion yield and productivity of permeabilized cells for 13‐HODE production were higher than those of purified and crude enzymes as well as nonpermeabilized cells. Therefore, permeabilized cells were efficient biocatalysts for 13‐HODE production. To the best of our knowledge, this is the first report of the production of 13‐HODE using cells.     

Alpha‐Lipoic Acid Supplementation Reduces mTORC1 Signaling in Skeletal Muscle from High Fat Fed,Obese Zucker Rats

The mammalian target of rapamycin (mTOR) signaling pathway is hyperactive in liver, adipose and skeletal muscle tissues of obese rodents. Alpha‐lipoic acid (αLA) has been well accepted as a weight‐loss treatment, though there are limited studies on its effect on mTOR signaling in high‐fat fed, obese rodents. Therefore, the goal of this study was to determine mTOR signaling and oxidative protein alterations in skeletal muscle of high‐fat fed, obese rats after αLA supplementation. Phosphorylation of the mTOR substrate, eukaryotic initiation factor (eIF) 4E‐binding protein 1 (4E‐BP1) and eIF4B were significantly reduced (p < 0.05) in muscle from αLA supplemented rats. Activation of AMP‐activated protein kinase (AMPK), an mTOR inhibitory kinase, was higher (p < 0.05) in the αLA group. Protein expression of markers of oxidative metabolism, acetyl CoA carboxylase (ACC), cytochrome c oxidase IV (COX IV), peroxisome proliferator‐activated receptor (PPAR), and PPAR gamma coactivator 1‐alpha (PGC‐1α) were significantly higher (p < 0.05) after αLA supplementation compared to non‐supplemented group. Our findings show that αLA supplementation limits the negative ramifications of consuming a high fat diet on skeletal muscle markers of oxidative metabolism and mTORC1 signaling.     

9‐Oxo‐10(E),12(Z),15(Z)‐Octadecatrienoic Acid Activates Peroxisome Proliferator‐Activated Receptor α in Hepatocytes

The peroxisome proliferator‐activated receptor (PPAR)α is mainly expressed in the liver and plays an important role in the regulation of lipid metabolism. It has been reported that PPARα activation enhances fatty acid oxidation and reduces fat storage. Therefore, PPARα agonists are used to treat dyslipidemia. In the present study, we found that 9‐oxo‐10(E),12(Z),15(Z)‐octadecatrienoic acid (9‐oxo‐OTA), which is a α‐linolenic acid (ALA) derivative, is present in tomato (Solanum lycopersicum) extract. We showed that 9‐oxo‐OTA activated PPARα and induced the mRNA expression of PPARα target genes in murine primary hepatocytes. These effects promoted fatty acid uptake and the secretion of β‐hydroxybutyrate, which is one of the endogenous ketone bodies. We also demonstrated that these effects of 9‐oxo‐OTA were not observed in PPARα‐knockout (KO) primary hepatocytes. To our knowledge, this is the first study to report that 9‐oxo‐OTA promotes fatty acid metabolism via PPARα activation and discuss its potential as a valuable food‐derived compound for use in the management of dyslipidemia.     

Derivation of a Retinoid X Receptor Scaffold from Peroxisome Proliferator‐Activated Receptor γ Ligand 1‐Di(1H‐indol‐3‐yl)methyl‐4‐trifluoromethylbenzene

PPARγ agonist DIM‐Ph‐4‐CF ₃ , a template for RXRα agonist (E)‐3‐[5‐di(1‐methyl‐1H‐indol‐3‐yl)methyl‐2‐thienyl] acrylic acid: DIM‐Ph‐CF₃ is reported to inhibit cancer growth independent of PPARγ and to interact with NR4A1. As both receptors dimerize with RXR, and natural PPARγ ligands activate RXR, DIM‐Ph‐4‐CF₃ was investigated as an RXR ligand. It displaces 9‐cis‐retinoic acid from RXRα but does not activate RXRα. Structure‐based direct design led to an RXRα agonist.

     

Branched‐chain fatty acids as activators of peroxisome proliferator‐activated receptors

We observed earlier that phytanic acid activated subtype α of the peroxisome proliferator‐activated receptor (PPAR) via the cytosolic liver‐type fatty acid‐binding protein (L‐FABP). In a further search for minor lipid nutrients that interact with genes, we explored here the potential of branched‐chain fatty acids to serve as agonists for the PPAR subtypes α, β and γ in rodent and human molecular test systems. Beyond chlorophyll‐derived pristanic and phytanic acids, bacteria‐derived iso‐ and anteiso‐fatty acids and avian‐derived ‘uropygial’ fatty acids were tested by transactivation assay. In addition, we studied binding of these fatty acids to recombinantly expressed PPAR ligand binding domains (LBDs) and to L‐FABP by competition with fluorescent parinaric acid. In contrast to single methyl‐branched agonists, multi methyl‐branched fatty acids had high transactivation potentials in either test system; in addition, some agonists of the latter were highly subtype selective. Multi methyl‐branched chain fatty acids were superior activators of human PPARγ, a preference not seen in the murine test system. High‐affinity binding of isoprenoid‐derived pristanic and phytanic acids to PPARγ‐LBD and to L‐FABP was observed, and also of pristanic acid to PPARα‐LBD. Polyketidic uropygial fatty acids bound to PPARγ‐LBD only, though weakly. As both isoprenoid and polyketidic fatty acids showed high activation potentials, it became clear that binding data determined in vitro cannot predict biological activity as determined by transactivation assay. For pristanic acid, however, a signalling path similar to that found for phytanic acid can be concluded. Taken together, multi methyl‐branched fatty acids of the human food chain can affect cellular metabolism through regulating gene expression.     

Responses of MAC‐T Cells to Inhibited Stearoyl‐CoA Desaturase 1 during cis‐9, trans‐11 Conjugated Linoleic Acid Synthesis

Cis‐9‐conjugated, trans‐11‐conjugated linoleic acid (CLA) is known for its positive activities on human health. The synthesis of cis‐9, trans‐11 CLA in mammary glands is generally thought to be catalyzed by stearoyl‐CoA desaturase 1 (SCD1), but this has not been rigorously established. In this study, we hypothesized that the inhibition of SCD1 (by CAY10566) would block the synthesis of cis‐9, trans‐11 CLA in bovine mammary alveolar cells (MAC‐T) cells. Results showed that MAC‐T cells incubated with 10 nM CAY10566 for 12 h (CAY) produced less cis‐9, trans‐11 CLA (p < 0.01), lower 14:1/(14:1 + 14:0)% (p < 0.01), more trans‐11 18:1 (TVA) accumulation (p < 0.01), and reduced SCD1 mRNA levels (p < 0.01) compared with the control group (CON). Moreover, the mRNA abundances of sterol regulatory element‐binding protein 1 [SREBPF1], acyl‐CoA synthetase short‐chain family member 2 [ACSS2], and lipin 1 [LPIN1] were significantly elevated when SCD1 was inhibited in the CAY group (p < 0.05). Taken together, CAY10566 inhibition of SCD1 resulted in lower cis‐9, trans‐11 CLA synthesis ability, and SREBF1, ACSSS2, and LPIN1 were negatively associated with SCD1. These findings not only provide the direct evidence that cis‐9, trans‐11 CLA synthesis is catalyzed by SCD1, but also help us understand the responses of MAC‐T cells to SCD1 inhibition.     

13-Hydroxy-9Z,15Z-Octadecadienoic Acid Production by Recombinant Cells Expressing Lactobacillus acidophilus 13-Hydratase

Recombinant Escherichia coli cells expressing linoleate 13‐hydratase from Lactobacillus acidophilus were permeabilized by treating with 0.2 M NaCl. The optimal conditions for the production of 13‐hydroxy‐9,15(Z,Z)‐octadecadienoic acid (13‐HODE) from α‐linolenic acid by permeabilized recombinant cells were pH 6.0, 40 °C, 7.5 % (v/v) methanol, 60 g/l permeabilized cells, and 15 g/l α‐linolenic acid. Under these conditions, permeabilized cells produced 7.5 g/l 13‐HODE after 6 h, with a conversion yield of 50 % (w/w) and a volumetric productivity of 1.25 g/l/h. These values were 161 and 160 % of those obtained by nonpermeabilized cells, respectively. To the best of our knowledge, this is the first report on the process optimization for the biotechnological production of 13‐HODE.     

A poly(γ‐glutamic acid)‐based hydrogel loaded with superoxide dismutase for wound healing

The purpose of this study was to develop a poly(γ‐glutamic acid) (γ‐PGA)‐based hydrogel loaded with superoxide dismutase (SOD) to accelerate wound healing. First, γ‐PGA was modified with taurine (γ‐PGAS), and then the SOD‐loaded γ‐PGAS/γ‐PGA hydrogel (SOD‐PGAS/PGA‐H) was prepared by cross‐linking of ethylene glycol diglycidyl ether. The swelling behavior and water vapor transmission rate revealed that PGAS/PGA‐H could create a moist environment for wound surface. In vitro kinetics of SOD release showed that SOD released from PGAS/PGA‐H maintained high activity and SOD‐PGAS/PGA‐H effectively scavenged the superoxide anion. The results of our fibroblast proliferation experiments showed that PGAS/PGA‐H had good cytocompatibility. The effects of SOD‐PGAS/PGA‐H on wound healing were examined in a Type I diabetic rat model with full‐thickness wounds. Twenty‐one days after grafted to wounds, SOD‐PGAS/PGA‐H exhibited a higher rate of wound healing than control group and showed increased collagen deposition and epithelialization. SOD‐PGAS/PGA‐H seems to promote better wound healing and thus might be a promising candidate for wound healing management. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42033.     

An increase in dietary n-3 fatty acids decreases a marker of bone resorption in humans

Human, animal, and in vitro research indicates a beneficial effect of appropriate amounts of omega-3 (n-3) polyunsaturated fatty acids (PUFA) on bone health. This is the first controlled feeding study in humans to evaluate the effect of dietary plant-derived n-3 PUFA on bone turnover, assessed by serum concentrations of N-telopeptides (NTx) and bone-specific alkaline phosphatase (BSAP). Subjects (n = 23) consumed each diet for 6 weeks in a randomized, 3-period crossover design: 1) Average American Diet (AAD; [34% total fat, 13% saturated fatty acids (SFA), 13% monounsaturated fatty acids (MUFA), 9% PUFA (7.7% LA, 0.8% ALA)]), 2) Linoleic Acid Diet (LA; [37% total fat, 9% SFA, 12% MUFA, 16% PUFA (12.6% LA, 3.6% ALA)]), and 3) α-Linolenic Acid Diet (ALA; [38% total fat, 8% SFA, 12% MUFA, 17% PUFA (10.5% LA, 6.5% ALA)]). Walnuts and flaxseed oil were the predominant sources of ALA. NTx levels were significantly lower following the ALA diet (13.20 ± 1.21 nM BCE), relative to the AAD (15.59 ± 1.21 nM BCE) (p < 0.05). Mean NTx level following the LA diet was 13.80 ± 1.21 nM BCE. There was no change in levels of BSAP across the three diets. Concentrations of NTx were positively correlated with the pro-inflammatory cytokine TNFα for all three diets. The results indicate that plant sources of dietary n-3 PUFA may have a protective effect on bone metabolism via a decrease in bone resorption in the presence of consistent levels of bone formation.     

Forms of n‐3 (ALA,C18:3n‐3 or DHA,C22:6n‐3) Fatty Acids Affect Carcass Yield,Blood Lipids,Muscle n‐3 Fatty Acids and Liver Gene Expression in Lambs

The effects of supplementing diets with n‐3 alpha‐linolenic acid (ALA) and docosahexaenoic acid (DHA) on plasma metabolites, carcass yield, muscle n‐3 fatty acids and liver messenger RNA (mRNA) in lambs were investigated. Lambs (n = 120) were stratified to 12 groups based on body weight (35 ± 3.1 kg), and within groups randomly allocated to four dietary treatments: basal diet (BAS), BAS with 10.7 % flaxseed supplement (Flax), BAS with 1.8 % algae supplement (DHA), BAS with Flax and DHA (FlaxDHA). Lambs were fed for 56 days. Blood samples were collected on day 0 and day 56, and plasma analysed for insulin and lipids. Lambs were slaughtered, and carcass traits measured. At 30 min and 24 h, liver and muscle samples, respectively, were collected for determination of mRNA (FADS1, FADS2, CPT1A, ACOX1) and fatty acid composition. Lambs fed Flax had higher plasma triacylglycerol, body weight, body fat and carcass yield compared with the BAS group (P < 0.001). DHA supplementation increased carcass yield and muscle DHA while lowering plasma insulin compared with the BAS diet (P < 0.01). Flax treatment increased (P < 0.001) muscle ALA concentration, while DHA treatment increased (P < 0.001) muscle DHA concentration. Liver mRNA FADS2 was higher and CPT1A lower in the DHA group (P < 0.05). The FlaxDHA diet had additive effects, including higher FADS1 and ACOX1 mRNA than for the Flax or DHA diet. In summary, supplementation with ALA or DHA modulated plasma metabolites, muscle DHA, body fat and liver gene expression differently.     

In situ injectable poly(γ‐glutamic acid) based biohydrogel formed by enzymatic crosslinking

Enzymatic crosslinking was developed to prepare in situ forming poly(γ‐glutamic acid) (γ‐PGA) based hydrogel in this study. First, the precursor of poly(γ‐glutamic acid)–tyramine (γ‐PGA–Ty) was synthesized through the reaction of carboxyl groups from a γ‐PGA backbone with tyramine. The structure of the grafted precursor was confirmed by ¹H‐NMR and Fourier transform infrared spectroscopy. After that, the crosslinking of the phenol‐containing γ‐PGA–Ty precursor was triggered by horseradish peroxidase in the presence of H₂O₂; this resulted in the formation of the γ‐PGA–Ty hydrogels. The equilibrium water content, morphology, enzymatic degradation rate, and mechanical properties of the hydrogels were characterized in detail. The data revealed that the well‐interconnected hydrogels had tunable water contents, mechanical properties, and degradability through adjustments of the composition. Furthermore, cell experiments proved the biocompatibility of the hydrogels by 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide assay. These characteristics provide an opportunity for the in situ formation of injectable biohydrogels as potential candidates in cell encapsulation and drug delivery. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42301.     

Synthesis and Biological Evaluation of N‐Substituted Sophocarpinic Acid Derivatives as Coxsackievirus B3 Inhibitors

A series of novel N‐substituted sophocarpinic acid derivatives was designed, synthesized, and evaluated for their anti‐enteroviral activities against coxsackievirus type B3 (CVB3) and coxsackievirus type B6 (CVB6) in Vero cells. Structure–activity relationship analysis revealed that the introduction of a benzenesulfonyl moiety on the 12‐nitrogen atom in (E)‐β,γ‐sophocarpinic acid might significantly enhance anti‐CVB3 activity. Among the derivatives, (E)‐12‐N‐(m‐cyanobenzenesulfonyl)‐β,γ‐sophocarpinic acid ( 11 m ), possessing a meta‐cyanobenzenesulfonyl group, exhibited potent activity against CVB3 with a selectivity index (SI) of 107. Furthermore, compound 11 m also showed a good oral pharmacokinetic profile, with an AUC value of 7.29 μM h^?1 in rats, and good safety through the oral route in mice, with an LD₅₀ value of >1000 mg kg^?1; these values suggest a druggable characteristic. Therefore, compound 11 m was selected for further investigation as a promising CVB3 inhibitor. We consider (E)‐β,γ‐N‐(benzenesulfonyl)sophocarpinic acids to be a novel class of anti‐CVB3 agents.     

Asymmetric Sharpless epoxidation of 13S‐hydroxy‐ 9Z, 11E‐octadecadienoic acid (13S‐HODE)

The asymmetric Sharpless epoxidation of methyl 13S‐hydroxy‐9Z, 11E‐octadeca‐dienoate (13S‐HODE, 1 ) with tert‐butyl hydroperoxide (TBHP) catalysed by titanium tetraisopropoxide {Ti(ⁱOPr)₄} in the presence of L(+)‐diisopropyl tartrate (L‐DIPT) gave methyl 13S‐hydroxy‐11S, 12S‐epoxy‐9Z‐octadecenoate 2 (erythro isomer) in 84% diastereomeric excess (de). The epoxidation of 1 with TBHP catalysed by Ti(ⁱOPr)₄ in the presence of D(‐)‐DIPT yielded methyl 13S‐hydroxy‐11RR12R‐epoxy‐9Z‐octadecenoate (threo isomer) 3 in 76% de.     

Development of 3‐Phenyl‐N‐(2‐(3‐phenylureido)ethyl)‐thiophene‐2‐sulfonamide Compounds as Inhibitors of Antiapoptotic Bcl‐2 Family Proteins

Antiapoptotic Bcl‐2 family proteins, such as Bcl‐x_L, Bcl‐2, and Mcl‐1, are often overexpressed in tumor cells, which contributes to tumor cell resistance to chemotherapies and radiotherapies. Inhibitors of these proteins thus have potential applications in cancer treatment. We discovered, through structure‐based virtual screening, a lead compound with micromolar binding affinity to Mcl‐1 (inhibition constant (K_i)=3 μM ). It contains a phenyltetrazole and a hydrazinecarbothioamide moiety, and it represents a structural scaffold not observed among known Bcl‐2 inhibitors. This work presents the structural optimization of this lead compound. By following the scaffold‐hopping strategy, we have designed and synthesized a total of 82 compounds in three sets. All of the compounds were evaluated in a fluorescence‐polarization binding assay to measure their binding affinities to Bcl‐x_L, Bcl‐2, and Mcl‐1. Some of the compounds with a 3‐phenylthiophene‐2‐sulfonamide core moiety showed sub‐micromolar binding affinities to Mcl‐1 (K_i=0.3–0.4 μM ) or Bcl‐2 (K_i≈1 μM ). They also showed obvious cytotoxicity on tumor cells (IC₅₀<10 μM ). Two‐dimensional heteronuclear single quantum coherence NMR spectra of three selected compounds, that is, YCW‐E5, YCW‐E10, and YCW‐E11, indicated that they bind to the BH3‐binding groove on Bcl‐x_L in a similar mode to ABT‐737. Several apoptotic assays conducted on HL‐60 cells demonstrated that these compounds are able to induce cell apoptosis through the mitochondrial pathway. We propose that the compounds with the 3‐phenylthiophene‐2‐sulfonamide core moiety are worth further optimization as effective apoptosis inducers with an interesting selectivity towards Mcl‐1 and Bcl‐2.     

Thermally crosslinked anionic hydrogels composed of poly(vinyl alcohol) and poly(γ‐glutamic acid): Preparation,characterization, and drug permeation behavior

pH‐sensitive anionic hydrogels composed of poly(vinyl alcohol) (PVA) and poly(γ‐glutamic acid) (γ‐PGA) were prepared by the freeze drying method and thermally crosslinked to suppress hydrogel deformation in water. The physical properties, swelling, and drug‐diffusion behaviors were characterized for the hydrogels. In the equilibrium swelling study, PVA/γ‐PGA hydrogels shrunk in pH regions below the pK_a (2.27) of γ‐PGA, whereas they swelled above the pK_a. In the drug‐diffusion study, the drug permeation rates of the PVA/γ‐PGA hydrogels were directly proportional to their swelling behaviors. The cytocompatibility test showed no cytotoxicity of the PVA/γ‐PGA hydrogels for the 3T3 fibroblast cell lines. The results of these studies suggest that hydrogels prepared from PVA and γ‐PGA could be used as orally administrable drug‐delivery systems. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2008     

Tricycloundecane Derivatives as Potential N‐Methyl‐D‐aspartate (NMDA) Receptor and Voltage‐Gated Calcium Channel Modulators

Neurodegenerative disorders are debilitating conditions characterised by progressive dysfunction and death of neuronal cells. Amidst the proposed mechanisms of neurodegeneration, the effects of excitotoxicity via N‐methyl‐D ‐aspartate (NMDA) receptor stimulation and activation of voltage‐gated calcium channels (VGCC) on neuronal cells are prominent. This has led to the development of polycyclic cage molecules such as NGP1‐01, which exhibit neuroprotective properties through NMDA receptor and VGCC modulation. The medicinal potential of structurally related tricycloundecanes that are open‐cage or rearranged polycyclic moieties has not been explored. This study is therefore focused on the synthesis of a series of novel tricycloundecane derivatives and their ability to inhibit NMDA receptors and VGCC. Significant NMDA receptor inhibition was observed for tricyclo[6.2.1.0^2,7]undec‐9‐ene‐3,6‐dione ( 4 , 78 %) and 6‐hydroxytricyclo[6.2.1.0^2,7]undec‐9‐en‐3‐one ( 5 , >95 %) at a concentration of 100 μM . The highest inhibitory activity was observed for 6‐(benzylimino)tricyclo[6.2.1.0^2,7]undec‐9‐en‐3‐one ( 9 , >95 %), which is in the same range as the inhibitory activity of MK‐801 (dizocilpine). In the VGCC inhibition assay, 6‐(benzylamino)tricyclo[6.2.1.0^2,7]undeca‐4,9‐dien‐3‐one ( 8 , 34 %), 9 (38 %) and 2‐(benzylamino)‐3,6‐epoxytricyclo[6.2.1.0^5,10]undecan‐9‐ol ( 12 , 40 %) showed statistically significant (p<0.05) VGCC inhibition.     

Organocatalytic Asymmetric Aldol Reaction of Ketones with β,γ‐Unsaturated α‐Keto Esters: An Efficient Access to Chiral Tertiary Alcohol Skeletons

This update describes a highly efficient organocatalytic aldol reaction of ketones and β,γ‐unsaturated α‐keto esters for constructing the chiral tertiary alcohol motif. With the application of 9‐amino(9‐deoxy)epi‐Cinchona alkaloid and an acidic additive as catalysts, both acyclic and cyclic ketones react with β,γ‐unsaturated α‐keto esters smoothly to afford aldol adducts in good to excellent yields and asymmetric induction. This protocol offers a new pathway for the construction of adjacent chiral carbon centers and the synthesis of chiral β‐hydroxy carbonyl compounds.