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1.
Gene ; 741: 144559, 2020 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-32169630

RESUMO

The fungi in order Mortierellales are attractive producers for long-chain polyunsaturated fatty acids (PUFAs). Here, the genome sequencing and assembly of a novel strain of Mortierella sp. BCC40632 were done, yielding 65 contigs spanning of 49,964,116 total bases with predicted 12,149 protein-coding genes. We focused on the acetyl-CoA in relevant to its derived metabolic pathways for biosynthesis of macromolecules with biological functions, including PUFAs, eicosanoids and carotenoids. By comparative genome analysis between Mortierellales and Mucorales, the signature genetic characteristics of the arachidonic acid-producing strains, including Δ5-desaturase and GLELO-like elongase, were also identified in the strain BCC40632. Remarkably, this fungal strain contained only n-6 pathway of PUFA biosynthesis due to the absence of Δ15-desaturase or ω3-desaturase gene in contrast to other Mortierella species. Four putative enzyme sequences in the eicosanoid biosynthetic pathways were identified in the strain BCC40632 and others Mortierellale fungi, but were not detected in the Mucorales. Another unique metabolic trait of the Mortierellales was the inability in carotenoid synthesis as a result of the lack of phytoene synthase and phytoene desaturase genes. The findings provide a perspective in strain optimization for production of tailored-made products with industrial applications.


Assuntos
Acetilcoenzima A/biossíntese , Ácido Araquidônico/genética , Genoma Fúngico/genética , Mortierella/metabolismo , Acetilcoenzima A/genética , Ácido Araquidônico/biossíntese , Vias Biossintéticas/genética , Ácidos Graxos Dessaturases/genética , Elongases de Ácidos Graxos/genética , Ácidos Graxos Insaturados/genética , Ácidos Graxos Insaturados/metabolismo , Mortierella/genética , Mucorales/genética , Mucorales/metabolismo , Ácido gama-Linolênico/genética , Ácido gama-Linolênico/metabolismo
2.
Enzyme Microb Technol ; 131: 109381, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31615662

RESUMO

Phytohormones are chemical messengers that have a positive effect at low concentrations on the biosynthesis of high-value compounds. Therefore, the effects of phytohormones on lipid and arachidonic acid (ARA) biosynthesis in Mortierella alpina were investigated in this study. At proper concentrations, the stimulatory effects of phytohormones on lipid production were determined to be as follows: 6-benzyl adenine (BA) > indole-3-acetic acid (IAA) > furfuryl adenine (KT) > gibberellin (GA) > indole-3-butyric acid (IBA) > abscisic acid (ABA). The results show that in the presence of 15 mg L-1 BA, the best positive effect was obtained, in which the lipid and ARA yields of M. alpina increased by 20.34% and 29.17%, respectively. Surprisingly, there was no synergy between the addition of two cytokinins (KT and BA), while adding cytokinins (KT or BA) and auxin (IAA) inhibited the growth of M. alpina and the ARA yield decreased by approximately 64%. Additional studies, such as those involving enzyme activity detection and quantitative real time polymerase chain reaction were carried out to check the fatty acid and lipid biosynthesis when the phytohormones were present. The activity of the main NADPH-supplying enzyme, 6-phosphoglucose dehydrogenase (G6PDH), increased by 19.52%. Moreover, the transcription levels of fatty acid synthase (FAS), Δ9-desaturase, and diacylglycerolacyltransferase (DGAT) increased by 9.3, 9.6 and 7.7 times, respectively, when only one type of phytohormone was present, indicating the enhancement of fatty acid and lipid biosynthesis in M. alpina. This study demonstrates the potential application of phytohormones for improving ARA yields of M. alpina.


Assuntos
Ácido Araquidônico/biossíntese , Vias Biossintéticas/efeitos dos fármacos , Mortierella/efeitos dos fármacos , Mortierella/metabolismo , Reguladores de Crescimento de Planta/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos
3.
J Agric Food Chem ; 67(35): 9851-9857, 2019 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-31418561

RESUMO

Arachidonic acid (ARA, C20:4) is a typical ω-6 polyunsaturated fatty acid with special functions. Using Yarrowia lipolytica as an unconventional chassis, we previously showed the performance of the Δ-6 pathway in ARA production. However, a significant increase in the Δ-9 pathway has rarely been reported. Herein, the Δ-9 pathway from Isochrysis galbana was constructed via pathway engineering, allowing us to synthesize ARA at 91.5 mg L-1. To further improve the ARA titer, novel enzyme fusions of Δ-9 elongase and Δ-8 desaturase were redesigned in special combinations containing different linkers. Finally, with the integrated pathway engineering and synthetic enzyme fusion, a 29% increase in the ARA titer, up to 118.1 mg/L, was achieved using the reconstructed strain RH-4 that harbors the rigid linker (GGGGS). The results show that the combined pathway and protein engineering can significantly facilitate applications of Y. lipolytica.


Assuntos
Ácido Araquidônico/biossíntese , Engenharia Metabólica , Yarrowia/genética , Yarrowia/metabolismo , Acetiltransferases/genética , Acetiltransferases/metabolismo , Ácidos Graxos Dessaturases/genética , Ácidos Graxos Dessaturases/metabolismo , Glucose/metabolismo , Haptófitas/enzimologia
4.
Insect Biochem Mol Biol ; 111: 103179, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31255640

RESUMO

Eicosanoids play crucial roles in mediating insect immune responses. In vertebrates, phospholipase A2 (PLA2) releases arachidonic acid (AA) from phospholipids (PLs) for biosynthesis of various eicosanoids. However, little AA is found in PLs of lepidopteran insects. Spodoptera exigua, a lepidopteran insect, is known to use eicosanoids to mediate immunity. Although AA was not detected in PLs of hemocytes and fat body (two immune tissues) of naïve larvae, it was detected at small but significant level after bacterial infection, suggesting induction of AA biosynthesis for immunity. Based on a mammalian AA biosynthetic pathway, this study hypothesizes that AA is synthesized from C18 polyunsaturated fatty acid (PUFA) precursor by subsequent desaturation and elongation reactions because PLs of S. exigua larvae are rich in linoleic acid. After inhibiting PLA2 activity to prevent release of free fatty acids, different PUFA precursors were injected to S. exigua larvae followed by assessment of eicosanoid-mediated cellular immune response. ω-6 PUFAs were effective in inducing immune response whereas α-linolenic acid (an ω-3 PUFA) was not. Several fatty acyl desaturases (SeDESs) have been predicted from S. exigua transcriptomes. Specific inhibitors against Δ5 or Δ6 DESs inhibited eicosanoid-mediated immune responses. Furthermore, RNA interference (RNAi) specific to Δ5 or Δ6 DES genes significantly suppressed eicosanoid-mediated immune responses. Four very long chain fatty acid elongase genes (SeEloV-A ∼ SeEloV-D) were predicted. Among respective RNAi treatments of these genes, only one RNAi treatment specific to type 5 elongase (SeEloV-B) suppressed eicosanoid-mediated immune response. These results suggest that S. exigua larvae can synthesize AA from linoleic acid via Δ5- and Δ6-desaturations by SeDESs along with chain elongation by SeEloV-B. Finally, this study showed significant fitness cost of uncontrolled AA biosynthesis. AA injection alone without bacterial challenge significantly induced both cellular and humoral immune responses. This unnecessary energy expense due to free AA resulted in reduced pupal size and decreased adult egg production. The detrimental effect of free AA explains physiological significance of little AA content in lepidopteran insects except for life-or-death situation such as pathogen infection.


Assuntos
Ácido Araquidônico/biossíntese , Spodoptera/metabolismo , Animais , Vias Biossintéticas , Eicosanoides/metabolismo , Escherichia coli , Ácidos Graxos Insaturados/metabolismo , Larva/genética , Larva/imunologia , Larva/metabolismo , Larva/microbiologia , Interferência de RNA , Spodoptera/genética , Spodoptera/imunologia , Spodoptera/microbiologia
5.
Nat Commun ; 10(1): 2602, 2019 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-31197136

RESUMO

Temperature is a key factor for determining the lifespan of both poikilotherms and homeotherms. It is believed that animals live longer at lower body temperatures. However, the precise mechanism remains largely unknown. Here, we report that autophagy serves as a boost mechanism for longevity at low temperature in the nematode Caenorhabditis elegans. The adiponectin receptor AdipoR2 homolog PAQR-2 signaling detects temperature drop and augments the biosynthesis of two ω-6 polyunsaturated fatty acids, γ-linolenic acid and arachidonic acid. These two polyunsaturated fatty acids in turn initiate autophagy in the epidermis, delaying an age-dependent decline in collagen contents, and extending the lifespan. Our findings reveal that the adiponectin receptor PAQR-2 signaling acts as a regulator linking low temperature with autophagy to extend lifespan, and suggest that such a mechanism may be evolutionally conserved among diverse organisms.


Assuntos
Autofagia/fisiologia , Proteínas de Caenorhabditis elegans/fisiologia , Caenorhabditis elegans/fisiologia , Longevidade/fisiologia , Proteínas de Membrana/fisiologia , Animais , Animais Geneticamente Modificados , Ácido Araquidônico/biossíntese , Temperatura Baixa , Colágeno/metabolismo , Epiderme/metabolismo , Ácidos Graxos Ômega-6/biossíntese , Interferência de RNA , Transdução de Sinais/fisiologia
6.
Biochim Biophys Acta Mol Cell Biol Lipids ; 1864(8): 1134-1144, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31048041

RESUMO

The interest in understanding the capacity of aquatic invertebrates to biosynthesise omega-3 (ω3) long-chain (≥C20) polyunsaturated fatty acids (LC-PUFA) has increased in recent years. Using the common octopus Octopus vulgaris as a model species, we previously characterised a ∆5 desaturase and two elongases (i.e. Elovl2/5 and Elovl4) involved in the biosynthesis of LC-PUFA in molluscs. The aim of this study was to characterise both molecularly and functionally, two methyl-end (or ωx) desaturases that have been long regarded to be absent in most animals. O. vulgaris possess two ωx desaturase genes encoding enzymes with ∆12 and ω3 regioselectivities enabling the de novo biosynthesis of the C18 PUFA 18:2ω6 (LA, linoleic acid) and 18:3ω3 (ALA, α-linolenic acid), generally regarded as dietary essential for animals. The O. vulgaris ∆12 desaturase ("ωx2") mediates the conversion of 18:1ω9 (oleic acid) into LA, and subsequently, the ω3 desaturase ("ωx1") catalyses the ∆15 desaturation from LA to ALA. Additionally, the O. vulgaris ω3 desaturase has ∆17 capacity towards a variety of C20 ω6 PUFA that are converted to their ω3 PUFA products. Particularly relevant was the affinity of the ω3 desaturase towards 20:4ω6 (ARA, arachidonic acid) to produce 20:5ω3 (EPA, eicosapentaenoic acid), as supported by yeast heterologous expression, and enzymatic activity exhibited in vivo when paralarvae were incubated in the presence of [1-14C]20:4ω6. These results confirmed that several routes enabling EPA biosynthesis are operative in O. vulgaris whereas ARA and docosahexaenoic acid (DHA, 22:6ω3) should be considered essential fatty acids since endogenous production appears to be limited.


Assuntos
Ácidos Graxos Dessaturases/metabolismo , Ácidos Graxos Insaturados/biossíntese , Octopodiformes/metabolismo , Animais , Ácido Araquidônico/biossíntese , Ácido Araquidônico/metabolismo , Ácidos Docosa-Hexaenoicos/biossíntese , Ácidos Docosa-Hexaenoicos/metabolismo , Ácido Eicosapentaenoico/biossíntese , Ácidos Graxos Dessaturases/genética , Ácido Linoleico/biossíntese , Octopodiformes/enzimologia , Ácido alfa-Linoleico/biossíntese
7.
Gene ; 706: 106-114, 2019 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-31039437

RESUMO

Biological significance of 18-carbon polyunsaturated fatty acids, γ-linolenic acid (GLA; C18:3 n-6) and dihomo-γ-linolenic acid (DGLA; C20:3 n-6) has gained much attention in the systematic development of optimized strains for industrial applications. In this work, a n-6 PUFAs-producing strain of Aspergillus oryzae was generated by manipulating metabolic reactions in fatty acid modification and triacylglycerol biosynthesis. The codon-optimized genes coding for Δ6-desaturase and Δ6-elongase of Pythium sp., and diacylglycerol acyltransferase 2 (mMaDGAT2) of Mortierella alpina were co-transformed in a single vector into A. oryzae BCC14614, yielding strain TD6E6-DGAT2. Comparative phenotypic analysis showed that a 70% increase of lipid titer was found in the engineered strain, which was a result of a significant increase in triacylglycerol (TAG) content (52.0 ±â€¯1.8% of total lipids), and corresponded to the increased size of lipid particles observed in the fungal cells. Interestingly, the proportions of GLA and DGLA in neutral lipids of the engineered strain were similar, with the highest titers obtained in the high C:N culture (29:0; 6% glucose) during the lipid-accumulating stage of growth. Time-course expression analysis of the engineered strain revealed transcriptional control of TAG biosynthesis through a co-operation between the native DGAT2 of A. oryzae and the transformed mMaDGAT2.


Assuntos
Aspergillus oryzae/metabolismo , Lipídeos/biossíntese , Engenharia Metabólica/métodos , Ácido 8,11,14-Eicosatrienoico/metabolismo , Ácido Araquidônico/biossíntese , Aspergillus oryzae/genética , Aspergillus oryzae/fisiologia , Vias Biossintéticas , Ácidos Graxos/metabolismo , Ácidos Graxos Insaturados/metabolismo , Proteínas Fúngicas/genética , Mortierella/genética , Triglicerídeos/biossíntese , Ácido gama-Linolênico/biossíntese
8.
Mar Biotechnol (NY) ; 21(1): 19-29, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30206714

RESUMO

As the first marine teleost demonstrated to have the ability of long-chain polyunsaturated fatty acids (LC-PUFA) biosynthesis from C18 PUFA precursors, the rabbitfish Siganus canaliculatus provides us a unique model for clarifying the regulatory mechanisms of LC-PUFA biosynthesis in teleosts aiming at the replacement of dietary fish oil (rich in LC-PUFA) with vegetable oils (rich in C18 PUFA precursors but devoid of LC-PUFA). In the study of transcription regulation of gene encoding the Δ6Δ5 fatty acyl desaturase (Δ6Δ5 Fads), a rate-limiting enzyme catalyzing the first step of LC-PUFA biosynthesis in rabbitfish, a binding site for the transcription factor (TF), peroxisome proliferator-activated receptor γ (Pparγ), was predicted in Δ6Δ5 fads2 promoter by bioinformatics analysis, and thus the present study focused on the regulatory roles of Pparγ on Δ6Δ5 fads2. First, the activity of the Δ6Δ5 fads2 promoter was proved to be downregulated by pparγ overexpression and upregulated by treatment of Pparγ antagonist (GW9662), respectively, in HEK 293T cells with the dual luciferase reporter assay. Pparγ was further confirmed to interact with the promoter by electrophoretic mobility shift assay. Moreover, in S. canaliculatus hepatocyte line (SCHL) cells, GW9662 decreased the expression of pparγ together with increase of Δ6Δ5 fads2 mRNA. Besides, Δ6Δ5 fads2 expression was increased by pparγ RNAi knockdown and reduced by its mRNA overexpression. Furthermore, knockdown of pparγ induced a high conversion of 18:3n-3 to 18:4n-3 and 18:2n-6 to 18:3n-6, while pparγ mRNA overexpression led to a lower conversion of that, and finally a significant decrease of 20:4n-6(ARA), 20:5n-3(EPA), and 22:6n-3(DHA) production. The results indicate that Pparγ is involved in the transcriptional regulation of liver LC-PUFA biosynthesis by targeting Δ6Δ5 fads2 in rabbitfish, which is the first report of Pparγ involvement in the regulation of LC-PUFA biosynthesis in teleosts.


Assuntos
Ácidos Graxos Dessaturases/genética , Proteínas de Peixes/genética , Peixes/genética , Fígado/metabolismo , PPAR gama/genética , Regiões Promotoras Genéticas , Anilidas/farmacologia , Animais , Organismos Aquáticos , Ácido Araquidônico/biossíntese , Ácido Araquidônico/genética , Sítios de Ligação , Linhagem Celular , Biologia Computacional , Ácidos Docosa-Hexaenoicos/biossíntese , Ácidos Docosa-Hexaenoicos/genética , Ácido Eicosapentaenoico/biossíntese , Ácido Eicosapentaenoico/genética , Ácidos Graxos Dessaturases/metabolismo , Proteínas de Peixes/metabolismo , Peixes/metabolismo , Regulação da Expressão Gênica , Genes Reporter , Células HEK293 , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Metabolismo dos Lipídeos/genética , Fígado/citologia , Luciferases/genética , Luciferases/metabolismo , PPAR gama/antagonistas & inibidores , PPAR gama/metabolismo , Ligação Proteica , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo
9.
BMC Biotechnol ; 18(1): 23, 2018 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-29716562

RESUMO

BACKGROUND: Arachidonic acid (ARA), which is a ω-6 polyunsaturated fatty acid, has a wide range of biological activities and is an essential component of cellular membranes in some human tissues. Mortierella alpina is the best strain for industrial production of ARA. To increase its yield of arachidonic acid, heavy ion beam irradiation mutagenesis of Mortierella alpina was carried out in combination with triclosan and octyl gallate treatment. RESULTS: The obtained mutant strain F-23 ultimately achieved an ARA yield of 5.26 g L- 1, which is 3.24 times higher than that of the wild-type strain. In addition, quantitative real-time PCR confirmed that the expression levels of fatty acid synthase (FAS), Δ5-desaturase, Δ6-desaturase, and Δ9-desaturase were all significantly up-regulated in the mutant F-23 strain, especially Δ6- and Δ9-desaturase, which were up-regulated 3- and 2-fold, respectively. CONCLUSIONS: This study confirmed a feasible mutagenesis breeding strategy for improving ARA production and provided a mutant of Mortierella alpina with high ARA yield.


Assuntos
Ácido Araquidônico/biossíntese , Mortierella/efeitos dos fármacos , Mortierella/efeitos da radiação , Reatores Biológicos , Ácidos Graxos Dessaturases/biossíntese , Ácido Graxo Sintases/biossíntese , Fermentação , Ácido Gálico/análogos & derivados , Ácido Gálico/farmacologia , Íons Pesados , Mortierella/genética , Mortierella/metabolismo , Mutagênese , Triclosan/farmacologia
10.
Appl Microbiol Biotechnol ; 102(13): 5763-5773, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29671003

RESUMO

This study systematically examined the effect of nitrogen and phosphorous stress on the formation of linoleic acid (LA), arachidonic acid (ARA), and eicosapentaenoic acid (EPA) in Porphyridium cruentum gy-h56. P. cruentum was cultivated in six different media conferring different conditions of nitrogen (N) sufficiency/deprivation and phosphorous (P) sufficiency/limitation/deprivation. Over a 16-day cultivation process, the dry-weight content, proportion of total fatty acids (TFAs), and the concentration in the medium of linoleic acid (LA) were greatly improved by a maximum of 2.5-, 1.6-, and 1.1-fold, respectively, under conditions of N or P deprivation compared with N and P sufficiency. In contrast, levels of EPA or ARA were not enhanced under N or P stress conditions. Additionally, the results showed that N deprivation weakened the impact of P deficiency on the content and proportions of LA and EPA, while P deprivation enhanced the impact of N starvation on the content and proportions of LA and EPA. The conditions of N sufficiency and P deprivation (N+P-) were the optimal conditions for the production of LA, while the optimal conditions for EPA, ARA, and TFAs production were N sufficiency and P limitation (N+P-lim). This study suggests the potential application of combining N removal from saline wastewater with the production of LA, ARA, EPA, and biodiesel.


Assuntos
Ácidos Graxos Insaturados/biossíntese , Microbiologia Industrial , Nitrogênio/metabolismo , Fósforo/metabolismo , Porphyridium/fisiologia , Estresse Fisiológico , Águas Residuárias/química , Ácido Araquidônico/biossíntese , Biocombustíveis , Ácido Eicosapentaenoico/biossíntese , Ácido Linoleico/biossíntese , Nitrogênio/isolamento & purificação , Nitrogênio/farmacologia , Fósforo/farmacologia , Porphyridium/efeitos dos fármacos
11.
Artigo em Inglês | MEDLINE | ID: mdl-29408384

RESUMO

Heavy and radioisotope labeling are commonly used methods to trace the pharmacological activity and metabolic fate of a biochemical or pharmaceutical in vivo. Recent years witnessed increased demand for molecules uniformly labeled with heavy carbon-13 (U-13C) or radioactive carbon-14 (U-14C) isotopes over singly labeled isotopic versions. Polyunsaturated fatty acids (PUFAs) represent one classic example where uniform 13C or 14C isotopic enrichment of the hydrocarbon backbone has numerous technical, metabolic and pharmacological advantages. PUFAs are usually produced in fungi or algae and uniform 13C or 14C enrichment of the hydrocarbon chain is achieved by feeding the microorganism a suitable U-13C or U-14C substrate. Previous literature methods describing the biosynthesis of U-13C or U-14C fatty acids reported variable isotopic enrichments that were less than anticipated and suffered from inconsistent growth of the microorganism due to radiotoxicity. In the present study, a single-tube method is described for the biosynthesis and extraction of U-13C and U-14C arachidonic acid (AA), a standard PUFA, from microcultures of the soil fungus Mortierella alpina. To produce U-13C-AA, a suspension of fungal spores and mycelial fragments was directly inoculated and grown into submerged cultures in a medium composed of U-13C-glucose and NaNO3 as the respective and only sources of carbon and nitrogen. The total 13C enrichment of AA was in excess of 95% and the percentage of U-13C-AA was in the range of 60-70%. These values have not been surpassed by previously reported methods. To produce U-14C-AA, the procedure was modified to limit the radiotoxic effects of 14C on fungal growth. Submerged cultures were initially grown on common 12C-glucose. Then, following glucose depletion, the biomass was collected and immediately cultured on U-14C-glucose. This approach is unprecedented in reported literature and has significantly limited the radiotoxic effects of 14C on the microorganism. Biomass transfer from 12C to 14C substrates was timed to keep an uninterrupted supply of carbon required to sustain the microorganism in the fatty acid synthesis mode and suppress ß-oxidation, a metabolic status that is prerequisite for enhanced isotopic purity of the 14C product. The specific activity of 14C enriched AA was estimated at 864 Ci/mol (range of 708-1020 Ci/mol) suggesting 69.2% (range of 56.7-81.7%) 14C enrichment along the AA hydrocarbon backbone. The present method used a single tube for microbial culture and lipid extraction to minimize manipulative losses and oxidative degradation of the labeled products. Production cost is comparatively cheaper to custom labeling and yields of U-13C and U-14C-AA are comparable to literature methods and sufficient for small scale in vitro and in vivo pharmacological studies.


Assuntos
Ácido Araquidônico/biossíntese , Isótopos de Carbono/química , Radioisótopos de Carbono/química , Mortierella/metabolismo , Ácido Araquidônico/isolamento & purificação , Técnicas de Cultura , Glucose/química
12.
Urolithiasis ; 46(2): 137-147, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28623397

RESUMO

Fatty acid (FA) composition of phospholipids in plasma and red blood cells (RBC) can influence calciuria, oxaluria and renal stone formation. In this regard, the ratio of arachidonic acid (AA) and its precursor linoleic acid (LA) appears to be important. Administration of γ-linolenic acid (GLA) has been shown to increase the concentration of dihomo-gamma linoleic acid (DGLA) relative to AA indicating that it may attenuate biosynthesis of the latter. Such effects have not been investigated in race groups having difference stone occurrence rates. Black (B) and white (W) healthy males ingested capsules containing linoleic acid (LA) and GLA, for 30 days. Plasma and RBC total phospholipid (TPL) FA profiles, serum and 24 h urine biomarkers of hypercalciuria and urinary stone risk factors were determined on days 0 and 30. Data were tested for statistical significance using GraphPadInstat version 3.02. Concentration and percentage content of DGLA in plasma TPL increased in W but not in B. Arachidonic acid (AA) did not change in either group. There was no change in calcium excretion in either group but oxalate and citrate excretion increased in W. We suggest that elongation of GLA to DGLA may occur more rapidly than desaturation of DGLA to AA in W and that depressed activity of the enzyme elongase may occur in B. Calciuric and citraturic effects may be dependent on the quantity of LA or on the mass ratio of LA/GLA in the FA supplement. Questions about the mooted DGLA-AA-oxaluria pathway arise. We speculate that there exists a potential for using GLA as a conservative treatment for hypocitraturia. The observation of different responses in B and W indicates that such differences may play a role in stone formation and prevention.


Assuntos
Hiperoxalúria/metabolismo , Redes e Vias Metabólicas/efeitos dos fármacos , Nefrolitíase/metabolismo , Fosfolipídeos/sangue , Ácido gama-Linolênico/uso terapêutico , Adulto , Ácido Araquidônico/biossíntese , Ácido Araquidônico/sangue , Biomarcadores/sangue , Biomarcadores/urina , Suplementos Nutricionais , Eritrócitos/metabolismo , Ácidos Graxos/sangue , Ácidos Graxos/metabolismo , Voluntários Saudáveis , Humanos , Hiperoxalúria/sangue , Hiperoxalúria/etnologia , Hiperoxalúria/urina , Ácidos Linoleicos/sangue , Ácidos Linoleicos/metabolismo , Masculino , Nefrolitíase/sangue , Nefrolitíase/etnologia , Nefrolitíase/urina , Fosfolipídeos/metabolismo , Projetos Piloto , Fatores de Risco , Adulto Jovem , Ácido gama-Linolênico/sangue , Ácido gama-Linolênico/metabolismo , Ácido gama-Linolênico/farmacologia
13.
Clin Nutr ; 37(3): 784-789, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-28716367

RESUMO

The essentiality of fatty acids was determined by the Burrs in the 1920s. It is commonly accepted that provision of linoleic (LA) and alpha-linolenic acids (ALA) prevents and reverses essential fatty acid deficiency (EFAD). Development of alternative injectable lipid emulsions (ILE) low in LA and ALA has raised concern about their ability to prevent EFAD. This review provides biochemical evidence coupled with observations from animal and human studies that aim to characterize which fatty acids are truly essential to prevent EFAD. Retroconversion pathways and mobilization from body stores suggest that arachidonic and docosahexaenoic acids (ARA and DHA - the main derivatives of LA and ALA, respectively) also prevent EFAD. Our group first proposed the essentiality of ARA and DHA by feeding mice exclusively these fatty acids and proving that they prevent EFAD. Survival for 5 generations on this diet provides additional evidence that growth and reproductive capabilities are maintained. Moreover, the use of fish oil-based ILE, with minimal LA and ALA and abundant DHA and ARA, for treatment of intestinal failure-associated liver disease, does not result in EFAD. These findings challenge the essentiality of LA and ALA in the presence of ARA and DHA. Evidence discussed in this review supports the idea that ARA and DHA can independently fulfill dietary essential fatty acid requirements. The imminent introduction of new ILE rich in ARA and DHA in the United States highlights the importance of understanding their essentiality, especially when provision of ALA and LA is below the established daily minimum requirement.


Assuntos
Emulsões Gordurosas Intravenosas/administração & dosagem , Ácidos Graxos Essenciais/administração & dosagem , Animais , Ácido Araquidônico/administração & dosagem , Ácido Araquidônico/biossíntese , Ácidos Docosa-Hexaenoicos/administração & dosagem , Ácidos Docosa-Hexaenoicos/biossíntese , Ácidos Graxos Essenciais/deficiência , Feminino , Óleos de Peixe/administração & dosagem , Humanos , Ácido Linoleico/administração & dosagem , Ácido Linoleico/metabolismo , Camundongos , Necessidades Nutricionais , Ratos , Estados Unidos , Ácido alfa-Linoleico/administração & dosagem , Ácido alfa-Linoleico/metabolismo
14.
Med Princ Pract ; 26(6): 561-566, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28898884

RESUMO

OBJECTIVE: To investigate free fatty acid levels and histopathological changes in the brain of rats fed a high fructose diet (HFrD) and to evaluate the effects of Mucuna pruriens, known to have antidiabetic activity, on these changes. MATERIALS AND METHODS: The study comprised 28 mature female Wistar rats. The rats were divided into 4 groups, each included 7 rats. Group 1: control; group 2: fed an HFrD; group 3: fed normal rat chow and M. pruriens; group 4: fed an HFrD and M. pruriens for 6 weeks. At the end of 6 weeks, the rats were decapitated, blood and brain tissues were obtained. Serum glucose and triglyceride levels were measured. Free fatty acid levels were measured in 1 cerebral hemisphere of each rat and histopathological changes in the other. The Mann-Whitney U test was used to compare quantitative continuous data between 2 independent groups, and the Kruskal-Wallis test was used to compare quantitative continuous data between more than 2 independent groups. RESULTS: Arachidonic acid and docosahexaenoic acid levels were significantly higher in group 2 than in group 1 (p < 0.05). Free arachidonic acid and docosahexaenoic acid levels in group 4 were significantly less than in group 2 (p < 0.05). Histopathological examination of group 2 revealed extensive gliosis, neuronal hydropic degeneration, and edema. In group 4, gliosis was much lighter than in group 2, and edema was not observed. Neuronal structures in group 4 were similar to those in group 1. CONCLUSIONS: The HFrD increased the levels of free arachidonic acid and docosahexaenoic acid probably due to membrane degradation resulting from possible oxidative stress and inflammation in the brain. The HFrD also caused extensive gliosis, neuronal hydropic degeneration, and edema. Hence, M. pruriens could have therapeutic effects on free fatty acid metabolism and local inflammatory responses in the brains of rats fed an HFrD.


Assuntos
Ácidos Graxos não Esterificados/biossíntese , Frutose/farmacologia , Mucuna , Extratos Vegetais/farmacologia , Animais , Ácido Araquidônico/biossíntese , Glicemia , Cérebro/efeitos dos fármacos , Cérebro/patologia , Ácidos Docosa-Hexaenoicos/biossíntese , Feminino , Gliose/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Wistar , Triglicerídeos/sangue
15.
J Ind Microbiol Biotechnol ; 44(8): 1225-1235, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28508118

RESUMO

To investigate the metabolic regulation against oxygen supply, comparative metabolomics was performed to explore the metabolic responses of Mortierella alpina in the process of arachidonic acid (ARA) production. More than 110 metabolites involved in Embden-Meyerhof-Parnas pathway, pentose phosphate pathway, tricarboxylic acid cycle, inositol phosphate metabolism, fatty acid biosynthesis, and amino acid metabolism were identified by gas chromatography-mass spectrometry. Samples at different aeration rates were clearly distinguished by principal components analysis and partial least squares analysis, indicating that oxygen supply had a profound effect on the metabolism of M. alpina. Eleven major metabolites were identified as potential biomarkers to be primarily responsible for the difference of metabolism. Further study of metabolic changes with the relevant pathways demonstrated that the levels of several intermediate metabolites in relation to central carbon metabolism changed remarkably via both processes and citrate and malate was supposed to play vital roles in polyunsaturated acid (PUFA) synthesis. Increase of myo-inositol and sorbitol were probably for osmo-regulation and redox balance, while enhanced phosphoric acid and pyroglutamic acid were supposed to have function in the activation of signal transduction pathway for stress resistance. The present study provides a novel insight into the metabolic responses of M. alpina to aeration rates and the metabolic characteristics during the ARA fermentation.


Assuntos
Ácido Araquidônico/biossíntese , Microbiologia Industrial , Metaboloma , Mortierella/metabolismo , Carbono/metabolismo , Fermentação , Cromatografia Gasosa-Espectrometria de Massas , Glicólise , Metabolômica , Análise de Componente Principal
16.
Bioresour Technol ; 235: 79-86, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28365352

RESUMO

To reduce fermentation-associated wastewater discharge and the cost of wastewater treatment, which further reduces the total cost of DHA and ARA production, this study first analyzed the composition of wastewater from Aurantiochytrium (DHA) and Mortierella alpina (ARA) fermentation, after which wastewater recycling technology for these fermentation processes was developed. No negative effects of DHA and ARA production were observed when the two fermentation wastewater methods were cross-recycled. DHA and ARA yields were significantly inhibited when the wastewater from the fermentation process was directly reused. In 5-L fed-batch fermentation experiments, using this cross-recycle technology, the DHA and ARA yields were 30.4 and 5.13gL-1, respectively, with no significant changes (P>0.05) compared to the control group, and the water consumption was reduced by half compared to the traditional process. Therefore, this technology has great potential in industrial fermentation for polyunsaturated fatty acid production.


Assuntos
Fermentação , Águas Residuárias , Ácido Araquidônico/biossíntese , Ácidos Graxos Insaturados , Mortierella
17.
Bioresour Technol ; 227: 142-146, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28013130

RESUMO

Arachidonic acid (ARA) is one of the most abundant polyunsaturated fatty acids (PUFAs) in the mammalian brain. Many enzymatically- and nonenzymatically-produced metabolic products have important and potent pharmacological properties. However, uniformly isotope labeled forms of ARA are not commercially available for studying the metabolic fates of ARA. This study describes a simple and efficient protocol for the biosynthesis of U-13C-ARA from U-13C-glucose, and U-14C-ARA from U-14C-glucose by Mortierella alpina. The protocols yield approximately 100nmol quantities of U-13C-ARA with an isotopic purity of 95% from a 500µl batch volume, and approximately 2µCi quantities of U-14C-ARA with an apparent specific activity in excess of 1200Ci/mol from a 250µl batch volume.


Assuntos
Ácido Araquidônico/biossíntese , Ácido Araquidônico/química , Isótopos de Carbono/química , Marcação por Isótopo/métodos , Mortierella/metabolismo , Radioisótopos de Carbono/química , Glucose/metabolismo
18.
Gene ; 591(2): 327-32, 2016 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-27265031

RESUMO

Nucleopolyhedroviruses (NPVs) is one group of Baculoviruses. The infection of NPV in silkworm is often lethal. To investigate the effective measures to stop the infection of NPV, we cloned cDNA encoding small heat shock protein 25.4 in Antheraea pernyi (Ap-HSP25.4). The translated amino acid sequence consisted of 223 residues with a calculated molecular mass of 25.4kDa and an isoelectronic point (pI) of 4.93. Quantitative real-time PCR was used to investigate the expression patterns and distribution profiles of Ap-sHSP25.4 before and after challenged with NPV. We found that the inhibitors of eicosanoid synthesis could suppress the transcription of Ap-sHSP25.4 in the fat body in a dose dependent manner. And arachidonic acid induced the expression of Ap-sHSP25.4. Thus, we concluded that sHSPs may be promising candidates to boost insect immunity in practice.


Assuntos
Proteínas de Choque Térmico Pequenas/biossíntese , Mariposas/virologia , /fisiologia , Animais , Ácido Araquidônico/biossíntese , Clonagem Molecular , Eicosanoides/metabolismo , Evolução Molecular , Genes de Insetos , Proteínas de Choque Térmico Pequenas/genética , Larva , Mariposas/genética , Mariposas/imunologia , Estresse Fisiológico
19.
Mol Biol Evol ; 33(7): 1726-39, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27188529

RESUMO

Long chain polyunsaturated fatty acids (LCPUFA) are bioactive components of membrane phospholipids and serve as substrates for signaling molecules. LCPUFA can be obtained directly from animal foods or synthesized endogenously from 18 carbon precursors via the FADS2 coded enzyme. Vegans rely almost exclusively on endogenous synthesis to generate LCPUFA and we hypothesized that an adaptive genetic polymorphism would confer advantage. The rs66698963 polymorphism, a 22-bp insertion-deletion within FADS2, is associated with basal FADS1 expression, and coordinated induction of FADS1 and FADS2 in vitro. Here, we determined rs66698963 genotype frequencies from 234 individuals of a primarily vegetarian Indian population and 311 individuals from the US. A much higher I/I genotype frequency was found in Indians (68%) than in the US (18%). Analysis using 1000 Genomes Project data confirmed our observation, revealing a global I/I genotype of 70% in South Asians, 53% in Africans, 29% in East Asians, and 17% in Europeans. Tests based on population divergence, site frequency spectrum, and long-range haplotype consistently point to positive selection encompassing rs66698963 in South Asian, African, and some East Asian populations. Basal plasma phospholipid arachidonic acid (ARA) status was 8% greater in I/I compared with D/D individuals. The biochemical pathway product-precursor difference, ARA minus linoleic acid, was 31% and 13% greater for I/I and I/D compared with D/D, respectively. This study is consistent with previous in vitro data suggesting that the insertion allele enhances n-6 LCPUFA synthesis and may confer an adaptive advantage in South Asians because of the traditional plant-based diet practice.


Assuntos
Ácido Araquidônico/biossíntese , Ácidos Graxos Dessaturases/genética , Seleção Genética , Adulto , Alelos , Ácido Araquidônico/genética , Ácido Araquidônico/metabolismo , Bases de Dados de Ácidos Nucleicos , Ácidos Graxos Dessaturases/metabolismo , Ácidos Graxos Insaturados/genética , Ácidos Graxos Insaturados/metabolismo , Feminino , Frequência do Gene/genética , Variação Genética , Haplótipos , Humanos , Mutação INDEL , Masculino , Fosfolipídeos/genética , Fosfolipídeos/metabolismo , Polimorfismo de Nucleotídeo Único , Adulto Jovem
20.
Bioprocess Biosyst Eng ; 39(7): 1129-36, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27004948

RESUMO

Polyunsaturated fatty acids (PUFAs) are highly appreciated on their nutritive value for human health and aquaculture. P. purpureum, one of the red microalgae acknowledged as a promising accumulator of ARA, was chosen as the target algae in the present research. Effects of sodium bicarbonate (0.04-1.2 g/L), temperature (25, 30 and 33 °C) and phosphate (0.00-0.14 g/L) on biomass yield, total fatty acids (TFA) and arachidonic acid (ARA) accumulation were investigated systemically. NaHCO3 dose of 0.8 g/L and moderate temperature of 30 °C were preferred. In addition, TFA and ARA production were significantly enhanced by an appropriate concentration of phosphate, and the highest TFA yield of 666.38 mg/L and ARA yield of 159.74 mg/L were obtained at a phosphate concentration of 0.035 g/L. Interestingly, with phosphate concentration continuing to fall, UFA/TFA and ARA/EPA ratios were increased accordingly, suggesting that phosphate limitation promoted unsaturated fatty acids and arachidonic acid biosynthesis. Low concentration of phosphate may be favored to increase the enzymatic activities of ∆6-desaturase, which played a key role in catalyzing the conversion of C16:0 to C18:2, and thus the selectivity of UFA increased. Meanwhile, the increase of ARA selectivity could be attributed to ω6 pathway promotion and ∆17-desaturase activity inhibition with phosphate limitation. Phosphate limitation strategy enhanced unsaturated fatty acids and ARA biosynthesis in P. purpureum, and can be applied in commercial scale manufacturing and commercialization of ARA.


Assuntos
Ácido Araquidônico/biossíntese , Ácidos Graxos Insaturados/metabolismo , Fosfatos/metabolismo , Porphyridium/metabolismo , Biomassa , Temperatura
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