RESUMO
Stonikacidin A (1), the first representative of a new class of 4-bromopyrrole alkaloids containing an aldonic acid core, was isolated from the marine sponge Lissodendoryx papillosa. The compound is named in honor of Prof. Valentin A. Stonik, who is one of the outstanding investigators in the field of marine natural chemistry. The structure of 1 was determined using NMR, MS analysis, and chemical correlations. The L-idonic acid core was established by the comparison of GC, NMR, MS, and optical rotation data of methyl-pentaacetyl-aldonates obtained from the hydrolysis products of 1 and standard hexoses. The L-form of the idonic acid residue in 1 was confirmed by GC analysis of pentaacetate of (S)-2-butyl ester of the hydrolysis product from 1 and compared with corresponding derivatives of L- and D-idonic acids. The biosynthetic pathway for stonikacidin A (1) was proposed. The alkaloid 1 inhibited the growth of Staphylococcus aureus and Escherichia coli test strains, as well as affected the formation of S. aureus and E. coli biofilms. Compound 1 inhibited the activity of sortase A. Molecular docking data showed that stonikacidin A (1) can bind with sortase A due to the interactions between its bromine atoms and some amino acid residues of the enzyme.
Assuntos
Alcaloides , Escherichia coli , Poríferos , Staphylococcus aureus , Animais , Poríferos/química , Staphylococcus aureus/efeitos dos fármacos , Alcaloides/farmacologia , Alcaloides/química , Alcaloides/isolamento & purificação , Escherichia coli/efeitos dos fármacos , Simulação de Acoplamento Molecular , Antibacterianos/farmacologia , Antibacterianos/química , Antibacterianos/isolamento & purificação , Pirróis/farmacologia , Pirróis/química , Pirróis/isolamento & purificação , Biofilmes/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Proteínas de Bactérias , Oceano Pacífico , Cisteína Endopeptidases , AminoaciltransferasesRESUMO
Ten new decalin polyketides, zosteropenilline M (1), 11-epi-8-hydroxyzosteropenilline M (2), zosteropenilline N (3), 8-hydroxyzosteropenilline G (4), zosteropenilline O (5), zosteropenilline P (6), zosteropenilline Q (7), 13-dehydroxypallidopenilline A (8), zosteropenilline R (9) and zosteropenilline S (10), together with known zosteropenillines G (11) and J (12), pallidopenilline A (13) and 1-acetylpallidopenilline A (14), were isolated from the ethyl acetate extract of the fungus Penicillium yezoense KMM 4679 associated with the seagrass Zostera marina. The structures of isolated compounds were established based on spectroscopic methods. The absolute configurations of zosteropenilline Q (7) and zosteropenilline S (10) were determined using a combination of the modified Mosher's method and ROESY data. The absolute configurations of zosteropenilline M (1) and zosteropenilline N (3) were determined using time-dependent density functional theory (TD-DFT) calculations of the ECD spectra. A biogenetic pathway for compounds 1-14 is proposed. The antimicrobial, cytotoxic and cytoprotective activities of the isolated compounds were also studied. The significant cytoprotective effects of the new zosteropenilline M and zosteropenillines O and R were found in a cobalt chloride (II) mimic in in vitro hypoxia in HEK-293 cells. 1-Acetylpallidopenilline A (14) exhibited high inhibition of human breast cancer MCF-7 cell colony formation with IC50 of 0.66 µM and its anticancer effect was reduced when MCF-7 cells were pretreated with 4-hydroxitamoxifen. Thus, we propose 1-acetylpallidopenilline A as a new xenoestrogen with significant activity against breast cancer.
Assuntos
Penicillium , Zosteraceae , Penicillium/química , Humanos , Linhagem Celular Tumoral , Policetídeos/farmacologia , Policetídeos/química , Policetídeos/isolamento & purificação , Células MCF-7 , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Organismos AquáticosRESUMO
Investigation of the Vietnamese marine sponge Rhabdastrella globostellata led to the isolation of two new polar isomalabaricanes: rhabdastrellosides A (1) and B (2). Their structures and stereochemistry were elucidated with the application of 1D and 2D NMR, HRESIMS, and HRESIMS/MS methods, as well as chemical modifications and GC-MS analysis. Metabolites 1 and 2 are the first isomalabaricanes with non-oxidized cyclopentane ring in the tricyclic core system. Moreover, having a 3-O-disaccharide moiety in their structures, they increase a very rare group of isomalabaricane glycosides. We report here a weak cytotoxicity of 1 and 2 toward human neuroblastoma SH-SY5Y cells and normal rat H9c2 cardiomyocytes, as well as the cytoprotective activity of rhabdastrelloside B (2) at 1 µM evaluated using CoCl2-treated SH-SY5Y and H9c2 cells.
Assuntos
Antineoplásicos , Neuroblastoma , Poríferos , Triterpenos , Animais , Humanos , Ratos , Estrutura Molecular , Glicosídeos/farmacologia , Glicosídeos/química , Ensaios de Seleção de Medicamentos Antitumorais , Triterpenos/química , Poríferos/química , Antineoplásicos/químicaRESUMO
Two new cyclopiane diterpenes and a new cladosporin precursor, together with four known related compounds, were isolated from the marine sediment-derived fungus Penicillium antarcticum KMM 4670, which was re-identified based on phylogenetic inference from ITS, BenA, CaM, and RPB2 gene regions. The absolute stereostructures of the isolated cyclopianes were determined using modified Mosher's method and quantum chemical calculations of the ECD spectra. The isolation from the natural source of two biosynthetic precursors of cladosporin from a natural source has been reported for the first time. The antimicrobial activities of the isolated compounds against Staphylococcus aureus, Escherichia coli, and Candida albicans as well as the inhibition of staphylococcal sortase A activity were investigated. Moreover, the cytotoxicity of the compounds to mammalian cardiomyocytes H9c2 was studied. As a result, new cyclopiane diterpene 13-epi-conidiogenone F was found to be a sortase A inhibitor and a promising anti-staphylococcal agent.
Assuntos
Diterpenos , Penicillium , Policetídeos , Animais , Estrutura Molecular , Policetídeos/farmacologia , Filogenia , Penicillium/química , Staphylococcus , Diterpenos/química , Sedimentos Geológicos , MamíferosRESUMO
New anthraquinone derivatives acruciquinones A-C (1-3), together with ten known metabolites, were isolated from the obligate marine fungus Asteromyces cruciatus KMM 4696. Acruciquinone C is the first member of anthraquinone derivatives with a 6/6/5 backbone. The structures of isolated compounds were established based on NMR and MS data. The absolute stereoconfigurations of new acruciquinones A-C were determined using ECD and quantum chemical calculations (TDDFT approach). A plausible biosynthetic pathway of the novel acruciquinone C was proposed. Compounds 1-4 and 6-13 showed a significant antimicrobial effects against Staphylococcus aureus growth, and acruciquinone A (1), dendryol B (4), coniothyrinone B (7), and ω-hydroxypachybasin (9) reduced the activity of a key staphylococcal enzyme, sortase A. Moreover, the compounds, excluding 4, inhibited urease activity. We studied the effects of anthraquinones 1, 4, 7, and 9 and coniothyrinone D (6) in an in vitro model of skin infection when HaCaT keratinocytes were cocultivated with S. aureus. Anthraquinones significantly reduce the negative impact of S. aureus on the viability, migration, and proliferation of infected HaCaT keratinocytes, and acruciquinone A (1) revealed the most pronounced effect.
Assuntos
Ascomicetos , Infecções Estafilocócicas , Staphylococcus aureus , Antraquinonas/farmacologiaRESUMO
The marine-derived fungal strains KMM 4718 and KMM 4747 isolated from sea urchin Scaphechinus mirabilis as a natural fungal complex were identified as Penicillium sajarovii and Aspergillus protuberus based on Internal Transcribed Spacer (ITS), partial ß-tubulin (BenA), and calmodulin (CaM) molecular markers as well as an ribosomal polymerase two, subunit two (RPB2) region for KMM 4747. From the ethyl acetate extract of the co-culture, two new polyketides, sajaroketides A (1) and B (2), together with (2'S)-7-hydroxy-2-(2'-hydroxypropyl)-5-methylchromone (3), altechromone A (4), norlichexanthone (5), griseoxanthone C (6), 1,3,5,6-tetrahydroxy-8-methylxanthone (7), griseofulvin (8), 6-O-desmethylgriseofulvin (9), dechlorogriseofulvin (10), and 5,6-dihydro-4-methyl-2H-pyran-2-one (11) were identified. The structures of the compounds were elucidated using spectroscopic analyses. The absolute configurations of the chiral centers of sajaroketides A and B were determined using time-dependent density functional theory (TDDFT)-based calculations of the Electronic Circular Dichroism (ECD) spectra. The inhibitory effects of these compounds on urease activity and the growth of Staphylococcus aureus, Escherichia coli, and Candida albicans were observed. Sajaroketide A, altechromone A, and griseofulvin showed significant cardioprotective effects in an in vitro model of S. aureus-induced infectious myocarditis.
Assuntos
Penicillium , Policetídeos , Staphylococcus aureus , Estrutura Molecular , Policetídeos/química , Griseofulvina/farmacologia , Fungos , Dicroísmo CircularRESUMO
The metabolic profile of the Aspergillus sp. 1901NT-1.2.2 sponge-associated fungal strain was investigated using the HPLC MS technique, and more than 23 peaks in the HPLC MS chromatogram were detected. Only two minor peaks were identified as endocrocin and terpene derivative MS data from the GNPS database. The main compound was isolated and identified as known anthraquinone derivative vismione E. The absolute stereochemistry of vismione E was established for the first time using ECD and quantum chemical methods. Vismione E showed high cytotoxic activity against human breast cancer MCF-7 cells, with an IC50 of 9.0 µM, in comparison with low toxicity for normal human breast MCF-10A cells, with an IC50 of 65.3 µM. It was found that vismione E inhibits MCF-7 cell proliferation and arrests the cell cycle in the G1 phase. Moreover, the negative influence of vismione E on MCF-7 cell migration was detected. Molecular docking of vismione E suggested the IMPDH2 enzyme as one of the molecular targets for this anthraquinone derivative.
Assuntos
Antineoplásicos , Poríferos , Animais , Humanos , Simulação de Acoplamento Molecular , Linhagem Celular Tumoral , Aspergillus , Fungos , Antineoplásicos/química , Antraquinonas/farmacologia , Estrutura MolecularRESUMO
This Special Issue was announced as a platform for authors studying the isolation and identification of various natural products with cytoprotective effects and those studying cytoprotective synthetic compounds [...].
Assuntos
Produtos Biológicos , Descoberta de Drogas , Produtos Biológicos/farmacologiaRESUMO
Toporosides A-D (1-4), new ω-glycosylated fatty acid amides, were isolated from the sponge Stelodoryx toporoki. The structures of these compounds, including absolute configurations of stereogenic centers, were established using analysis of 1D and 2D NMR, ECD, and HR mass spectra as well as chemical transformations. Toporosides A (1) and B (2) are the first lipids containing a cyclopentenyl α,ß-unsaturated carbonyl moiety in the polymethylene chain. Toporoside C (3) is likely a precursor, which undergoes intramolecular aldol condensation to produce 1 and 2. Toporosides A, C, and D showed protective effects against TNF-α-induced injury in H9c2 cardiomyocytes.
Assuntos
Amidas , Poríferos , Amidas/química , Animais , Ácidos Graxos/química , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Poríferos/químicaRESUMO
Marine sediment derived fungi are a very interesting source of biologically active compounds [...].
Assuntos
Produtos Biológicos , Fungos , Produtos Biológicos/farmacologia , Sedimentos Geológicos/microbiologiaRESUMO
Three new tripeptide derivatives asterripeptides A-C (1-3) were isolated from Vietnamese mangrove-derived fungus Aspergillus terreus LM.5.2. Structures of isolated compounds were determined by a combination of NMR and ESIMS techniques. The absolute configurations of all stereocenters were determined using the Murfey's method. The isolated compounds 1-3 contain a rare fungi cinnamic acid residue. The cytotoxicity of isolated compounds against several cancer cell lines and inhibition ability of sortase A from Staphylococcus aureus of asterripeptides A-C were investigated.
Assuntos
Antibacterianos/farmacologia , Antineoplásicos/farmacologia , Aspergillus , Magnoliopsida , Peptídeos/farmacologia , Animais , Antibacterianos/química , Antineoplásicos/química , Organismos Aquáticos , Linhagem Celular Tumoral , Humanos , Testes de Sensibilidade Microbiana , Peptídeos/química , Staphylococcus aureus/efeitos dos fármacosRESUMO
Chemical investigation of a coculture of the marine-derived fungi Beauveria felina KMM 4639 and Aspergillus carneus KMM 4638 led to the identification of three new drimane-type sesquiterpenes, asperflavinoids B, D and E (2, 4, 5), and nine previously reported related compounds. The structures of these compounds were established using spectroscopic methods and by comparison with known analogues. We also investigated the cytotoxic activity of the isolated compounds against several cancer and normal cell lines. Asperflavinoid C (3) and ustusolate E (9) exerted a significant effect on human breast cancer MCF-7 cell viability, with IC50 values of 10 µM, and induced in caspase-dependent apoptosis and arrest of the MCF-7 cell cycle in the G2/M phase in these cells.
Assuntos
Antineoplásicos , Beauveria , Sesquiterpenos , Antineoplásicos/química , Aspergillus , Beauveria/química , Linhagem Celular Tumoral , Técnicas de Cocultura , Humanos , Estrutura Molecular , Sesquiterpenos Policíclicos , Sesquiterpenos/químicaRESUMO
The new polyketides lopouzanones A and B, as well as the new 1-O-acetyl and 2-O-acetyl derivatives of dendrodochol B, were isolated from the sponge-derived marine fungus Lopadostoma pouzarii strain 168CLC-57.3. Moreover, six known polyketides, gliorosein, balticolid, dendrodolide G, dihydroisocoumarine, (-)-5-methylmellein, and dendrodochol B, were identified. The structures of the isolated compounds were determined by a combination of NMR and ESIMS techniques. The absolute configurations of the lopouzanones A and B were determined using the Mosher's method. The cytotoxicity of the isolated compounds against human prostate cancer cells PC-3 and normal rat cardiomyocytes H9c2 was investigated. Gliorosein showed weak DPPH radical-scavenging activity and in vitro cardioprotective effects toward rotenone toxicity and CoCl2-mimic hypoxia.
Assuntos
Ascomicetos , Policetídeos , Humanos , Ratos , Animais , Policetídeos/química , Ascomicetos/química , Espectroscopia de Ressonância Magnética/métodos , Estrutura MolecularRESUMO
Marine sediments are characterized by intense degradation of sedimenting organic matter in the water column and near surface sediments, combined with characteristically low temperatures and elevated pressures. Fungi are less represented in the microbial communities of sediments than bacteria and archaea and their relationships are competitive. This results in wide variety of secondary metabolites produced by marine sediment-derived fungi both for environmental adaptation and for interspecies interactions. Earlier marine fungal metabolites were investigated mainly for their antibacterial and antifungal activities, but now also as anticancer and cytoprotective drug candidates. This review aims to describe low-molecular-weight secondary metabolites of marine sediment-derived fungi in the context of their biological activity and covers research articles published between January 2016 and November 2020.
Assuntos
Anti-Infecciosos/farmacologia , Anti-Inflamatórios/farmacologia , Antineoplásicos/farmacologia , Fungos/metabolismo , Sedimentos Geológicos/microbiologia , Plâncton/efeitos dos fármacos , Metabolismo Secundário , Microbiologia da ÁguaRESUMO
Cerebrosides are glycosylated sphingolipids, and in mammals they contribute to the pro-/anti-inflammatory properties and innate antimicrobial activity of the skin and mucosal surfaces. Staphylococcus aureus infection can develop, not only from minor scratches of the skin, but this pathogen can also actively promote epithelial breach. The effect of cerebroside flavuside B from marine sediment-derived fungus Penicillium islandicum (Aniva Bay, the Sea of Okhotsk) on viability, apoptosis, total caspase activity, and cell cycle in human epidermal keratinocytes HaCaT line co-cultivated with S. aureus, as well as influence of flavuside B on LPS-treated HaCaT cells were studied. Influence of flavuside B on bacterial growth and biofilm formation of S. aureus and its effect on the enzymatic activity of sortase A was also investigated. It was found S. aureus co-cultivated with keratinocytes induces caspase-depended apoptosis and cell death, arrest cell cycle in the G0/G1 phase, and increases in cellular immune inflammation. Cerebroside flavuside B has demonstrated its antimicrobial and anti-inflammatory properties, substantially eliminating all the negative consequences caused by co-cultivation of keratinocytes with S. aureus or bacterial LPS. The dual action of flavuside B may be highly effective in the treatment of bacterial skin lesions and will be studied in the future in in vivo experiments.
Assuntos
Antibacterianos/farmacologia , Cerebrosídeos/farmacologia , Glicoesfingolipídeos/farmacologia , Queratinócitos/efeitos dos fármacos , Penicillium , Dermatopatias Bacterianas/prevenção & controle , Infecções Estafilocócicas/prevenção & controle , Staphylococcus aureus/efeitos dos fármacos , Organismos Aquáticos , Células HaCaT/efeitos dos fármacos , HumanosRESUMO
Seven new deoxyisoaustamide derivatives (1-7) together with known compounds (8-10) were isolated from the coral-derived fungus Penicillium dimorphosporum KMM 4689. Their structures were established using spectroscopic methods, X-ray diffraction analysis and by comparison with related known compounds. The absolute configurations of some alkaloids were determined based on CD and NOESY data as well as biogenetic considerations. The cytotoxic and neuroprotective activities of some of the isolated compounds were examined and structure-activity relationships were pointed out. New deoxyisoaustamides 4-6 at concentration of 1 µM revealed a statistical increase of PQ(paraquat)-treated Neuro-2a cell viability by 30-39%.
Assuntos
Antozoários , Antineoplásicos/isolamento & purificação , Indóis/isolamento & purificação , Penicillium/isolamento & purificação , Animais , Antozoários/química , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Cristalografia por Raios X/métodos , Humanos , Indóis/química , Indóis/farmacologia , Penicillium/químicaRESUMO
The influence of p-terphenyl polyketides 1-3 from Aspergillus candidus KMM 4676 and cerebroside flavuside B (4) from Penicillium islandicum (=Talaromyces islandicus) against the effect of neurotoxins, rotenone and paraquat, on Neuro-2a cell viability by MTT and LDH release assays and intracellular ROS level, as well as DPPH radical scavenging activity, was investigated. Pre-incubation with compounds significantly diminished the ROS level in rotenone- and paraquat-treated cells. It was shown that the investigated polyketides 1-3 significantly increased the viability of rotenone- and paraquat-treated cells in two of the used assays but they affected only the viability of paraquat-treated cells in the LDH release assay. Flavuside B statistically increased the viability of paraquat-treated cells in both MTT and LDH release assays, however, it increased the viability of rotenone-treated cells in the LDH release assay. Structure-activity relationships for p-terphenyl derivatives, as well as possible mechanisms of cytoprotective action of all studied compounds, were discussed.
Assuntos
Aspergillus/química , Citoproteção/efeitos dos fármacos , Glicoesfingolipídeos/farmacologia , Neuroblastoma/tratamento farmacológico , Fármacos Neuroprotetores/farmacologia , Neurotoxinas/toxicidade , Policetídeos/farmacologia , Animais , Linhagem Celular Tumoral , Sobrevivência Celular , Herbicidas/toxicidade , Inseticidas/toxicidade , Camundongos , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Fármacos Neuroprotetores/química , Paraquat/toxicidade , Policetídeos/química , Espécies Reativas de Oxigênio , Rotenona/toxicidadeRESUMO
Low molecular weight secondary metabolites of marine fungi Aspergillus flocculosus, Aspergillus terreus and Penicillium sp. from Van Phong and Nha Trang Bays (Vietnam) were studied and a number of polyketides, bis-indole quinones and terpenoids were isolated. The structures of the isolated compounds were determined by 1D and 2D NMR and HR-ESI-MS techniques. Stereochemistry of some compounds was established based on ECD data. A chemical structure of asterriquinone F (6) was thoroughly described for the first time. Anthraquinone (13) was firstly obtained from a natural source. Neuroprotective influences of the isolated compounds against 6-OHDA, paraquat and rotenone toxicity were investigated. 4-Hydroxyscytalone (1), 4-hydroxy-6-dehydroxyscytalone (2) and demethylcitreoviranol (3) have shown significant increasing of paraquat- and rotenone-treated Neuro-2a cell viability and anti-ROS activity.
Assuntos
Antiparkinsonianos/farmacologia , Aspergillus/metabolismo , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Doença de Parkinson/tratamento farmacológico , Penicillium/metabolismo , Animais , Antiparkinsonianos/isolamento & purificação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Camundongos , Estrutura Molecular , Neurônios/metabolismo , Neurônios/patologia , Fármacos Neuroprotetores/isolamento & purificação , Estresse Oxidativo/efeitos dos fármacos , Paraquat/toxicidade , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Espécies Reativas de Oxigênio/metabolismo , Rotenona/toxicidade , Metabolismo Secundário , Relação Estrutura-Atividade , VietnãRESUMO
The results of an investigation of the protective effects of five lanostane triterpenoids: 3ß-acetoxy-7ß,8ß-epoxy-5α-lanost-24-en-30,9α-olide (1), 3ß-hydroxy-7ß,8ß-epoxy-5α-lanost-24-en- 30,9α-olide (2), 29-nor-penasterone (3), penasterone (4), and acetylpenasterol (5), from a marine sponge, Penares sp., against paraquat-induced neuroblastoma Neuro-2a cell damage, are described. The influence of all compounds on viability of the Neuro-2a cells treated with paraquat (PQ) was studied with MTT and fluorescein diacetate assays as well as propidium iodide straining. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity of the compounds as well as their influence on reactive oxygen species (ROS) level and mitochondrial membrane potential in PQ-treated neuronal cells were analyzed. Finally, the effect of the compounds on intracellular level of heat shock protein 70 kDa (Hsp70) and neurite outgrowth in PQ-treated Neuro-2a cells were studied. Studied triterpenoids demonstrated protective effects against PQ-induced neurotoxicity associated with the ability to reduce ROS intracellular level and diminish mitochondrial dysfunction. Acetylpenasterol (5), as a more promising neuroprotective compound, significantly increased the viability of Neuro-2a cells incubated with PQ as well as decreased intracellular ROS level in these cells. Moreover, acetylpenasterol induced Hsp70 expression in PQ-treated cells. It was also shown to inhibit PQ-induced neurite loss and recovered the number of neurite-bearing cells. The relationship between neuroprotective activity of the investigated compounds 1-5 and their chemical structure was also discussed.
Assuntos
Metaboloma , Neurotoxinas/toxicidade , Paraquat/toxicidade , Poríferos/química , Triterpenos/metabolismo , Animais , Compostos de Bifenilo/química , Morte Celular/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sequestradores de Radicais Livres/farmacologia , Proteínas de Choque Térmico HSP70/metabolismo , Metaboloma/efeitos dos fármacos , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Crescimento Neuronal/efeitos dos fármacos , Picratos/química , Espécies Reativas de Oxigênio/metabolismo , Triterpenos/químicaRESUMO
Until 2004, the secondary metabolites of marine organisms of the Vietnamese territorial waters had been studied very poorly. Only four new compounds were isolated from 1977 to 2003. Joint Russian-Vietnamese expeditions aboard the research vessel 'Akademik Oparin' made it possible to study in detail the chemical diversity of marine micro- and macroorganisms. As a result of five expeditions, more than 250 low-molecular weight natural compounds, including 117 new metabolites, were isolated from marine invertebrates and microfilamentous fungi. Their biological activities, such as cytotoxic, cytoprotective, and antioxidant activities, were investigated. Information about the structure and biological activity of the compounds, the source for their isolation and the geographical location of the objects is summarized in this review.