Supplementation with rac-GR24 Facilitates the Accumulation of Biomass and Astaxanthin in Two Successive Stages of Haematococcus pluvialis Cultivation.

The unicellular freshwater green alga Haematococcus pluvialis has attracted much research attention due to its biosynthetic ability for large amounts of astaxanthin, a blood-red ketocarotenoid that is used in cosmetics, nutraceuticals, and pharmaceuticals. Recently, numerous studies have investigated the functions of natural astaxanthin; however, the high cost of the production of astaxanthin from H. pluvialis cultures restricts its commercial viability. There is an urgent need to fulfill commercial demands by increasing astaxanthin accumulation from H. pluvialis cultures. In this study, we discovered that treatment of H. pluvialis cultures at the beginning of the macrozooid stage (day 0) with 1 µM rac-GR24, a synthetic analogue of strigolactones (a class of phytohormones), led to significant increases in biomass [up to a maximum dry cell weight (DCW) of 0.53 g/L] during the macrozooid stage and astaxanthin (from 0.63 to 5.32% of DCW) during the hematocyst stage. We elucidated that this enhancement of biomass accumulation during the macrozooid stage by rac-GR24 is due to its increasing CO2 utilization efficiency in photosynthesis and carbohydrate biosynthesis. We also found that rac-GR24 stimulated the overproduction of nicotinamide adenine dinucleotide phosphate (NADPH) and antioxidant enzymes in H. pluvialis cultures, which alleviated the oxidative damage caused by reactive oxygen species generated during the hematocyst stage due to the exhaustion of nitrogen supplies. Moreover, rac-GR24 treatment of H. pluvialis synergistically altered the activity of the pathways of fatty acid biosynthesis and astaxanthin esterification, which resulted in larger amounts of astaxanthin being generated by rac-GR24-treated cultures than by controls. In summary, we have developed a feasible and economic rac-GR24-assisted strategy that increases the amounts of biomass and astaxanthin generated by H. pluvialis cultures, and have provided novel insights into the mechanistic roles of rac-GR24 to achieve these effects.

Hydrolysis of organophosphorus by diatom purple acid phosphatase and sequential regulation of cell metabolism.

Phosphorus (P) limitation affects phytoplankton growth and population size in aquatic systems, and consequently limits aquatic primary productivity. Plants have evolved a range of metabolic responses to cope with P limitation, such as accumulation of purple acid phosphatases (PAPs) to enhance acquisition of phosphates. However, it remains unknown whether algae have evolved a similar mechanism. In this study, we examined the role of PAPs in the model microalga Phaeodactylum tricornutum. Expression of PAP1 was enhanced in P. tricornutum cells grown on organophosphorus compared to inorganic phosphate. PAP1 overexpression improved cellular growth and biochemical composition in a growth-phase dependent manner. PAP1 promoted growth and photosynthesis during growth phases and reallocated carbon flux towards lipogenesis during the stationary phase. PAP1 was found to be localized in the endoplasmic reticulum and it orchestrated the expression of genes involved in key metabolic pathways and translocation of inorganic P (Pi), thereby improving energy use, reducing equivalents and antioxidant potential. RNAi of PAP1 induced expression of its homolog PAP2, thereby compensating for the Pi scavenging activity of PAP1. Our results demonstrate that PAP1 brings about sequential regulation of metabolism, and provide novel insights into algal phosphorus metabolism and aquatic primary productivity.

[Electroacupuncture combined with PNF on proprioception and motor function of lower limbs in stroke patients: a randomized controlled trial].

OBJECTIVE: To observe the clinical efficacy and correlation of electroacupuncture combined with proprioceptive neuromuscular facilitation (PNF) on proprioception and motor function of lower limbs in stroke patients. METHODS: A total of 96 stroke patients were randomized into an electroacupuncture (EA) group, a PNF group and a combination group, 32 cases in each one. In the EA group, acupuncture was applied at cephalic motor and sensory areas, Huantiao (GB 30), Yanglingquan (GB 34), Xuanzhong (GB 39), Zusanli (ST 36) and Sanyinjiao (SP 6) on affected side, and electroacupuncture was adopted at Yanglingquan (GB 34) and Xuanzhong (GB 39), continuous wave and 2 Hz in frequency for 20 min, once every day. In the PNF group, PNF was performed for 20 min, once a day. In the combination group, electroacupuncture was given before PNF, once a day. 4 weeks as one course and totally 3 courses were required, the effect was followed up after half a year. Before treatment, after 4, 8, 12 weeks of treatment and in follow-up, the average trace error (ATE) and Time, the scores of Fugl-Meyer scale (FMA) and modified Barthel index (MBI) were observed in the 3 groups. Correlation analysis between ATE, Time and FMA was performed. RESULTS: ① The total effective rate in the combination group was 90.3% (28/31), which was superior to 64.5% (20/31) in the EA group and 62.5% (20/32) in the PNF group (P<0.05). ②After 4, 8, 12 weeks of treatment and in follow-up, the ATE and Time in the 3 groups were reduced compared with before treatment (P<0.05, P<0.01). After 8, 12 weeks of treatment and in follow-up, the ATE in the combination group were lower than the EA group and the PNF group (P<0.05, P<0.01). After 12 weeks of treatment and in follow-up, the Time in the combination group were lower than the EA group and the PNF group (P<0.05, P<0.01). ③After 4, 8, 12 weeks of treatment and in follow-up, the FMA scores in the EA group and the combination group were increased compared with before treatment (P<0.01). After 8, 12 weeks of treatment and in follow-up, the FMA scores in the PNF group were increased compared with before treatment (P<0.01). After 8, 12 weeks of treatment and in follow-up, the FMA scores in the combination group were higher than the EA group and the PNF group (P<0.05, P<0.01). ④After 4, 8, 12 weeks of treatment and in follow-up, the MBI scores in the 3 groups were increased compared with before treatment (P<0.01). After 8, 12 weeks of treatment and in follow-up, the MBI scores in the combination group were higher than the EA group (P<0.01). After 12 weeks of treatment and in follow-up, the MBI scores in the combination group were higher than the PNF group (P<0.01). ⑤ The correlation coefficients of ATE, Time and FMA were from 0.4 to 0.75 (P<0.05), suggesting a moderate intensity correlation. CONCLUSION: Electroacupuncture, PNF and combination therapy can improve proprioception and motor function of lower limbs and activities of daily living, and combination therapy has a better effect. Proprioception and motor function have a strong correlation in the recovery of stroke patients.

Inhibition of five natural products from Chinese herbs on the growth of Chattonella marina.

The effects of five natural products from Chinese herbs including evodiamine, curcumin, 4-methoxysalicylaldehyde, esculin hydrate, and gramine on the growth of Chattonella marina, one of the most noxious red tide algae, were observed. Among them, gramine exhibited the highest inhibitory rate with LC50, 96h of 0.51 mg/l. After exposure to gramine, the activities of superoxide dismutase (SOD) and catalase (CAT), and content of malondialdehyde (MDA) increased in C. marina, suggesting that gramine could induce microalgae oxidative stress. In addition, chlorophyll a and the maximum quantum yield of photosynthesis (Fv/Fm) decreased following exposure to gramine, indicating the inhibition of photosynthesis activity in the microalgae. Combined with the fast inhibition against the algal cells and environmentally friendly character of gramine, we proposed that gramine might be a potential algaecide against marine harmful algae and that the oxidative damage and photosynthesis inhibition might be responsible for the toxicity of gramine on harmful algae.

Examination of metabolic responses to phosphorus limitation via proteomic analyses in the marine diatom Phaeodactylum tricornutum.

Phosphorus (P) is an essential macronutrient for the survival of marine phytoplankton. In the present study, phytoplankton response to phosphorus limitation was studied by proteomic profiling in diatom Phaeodactylum tricornutum in both cellular and molecular levels. A total of 42 non-redundant proteins were identified, among which 8 proteins were found to be upregulated and 34 proteins were downregulated. The results also showed that the proteins associated with inorganic phosphate uptake were downregulated, whereas the proteins involved in organic phosphorus uptake such as alkaline phosphatase were upregulated. The proteins involved in metabolic responses such as protein degradation, lipid accumulation and photorespiration were upregulated whereas energy metabolism, photosynthesis, amino acid and nucleic acid metabolism tend to be downregulated. Overall our results showed the changes in protein levels of P. tricornutum during phosphorus stress. This study preludes for understanding the role of phosphorous in marine biogeochemical cycles and phytoplankton response to phosphorous scarcity in ocean. It also provides insight into the succession of phytoplankton community, providing scientific basis for elucidating the mechanism of algal blooms.

Systems-level analysis of the metabolic responses of the diatom Phaeodactylum tricornutum to phosphorus stress.

Phosphorus is an important macronutrient. To understand the molecular and cellular responses to phosphorus stress better, transcriptome profiling in combination with biochemical investigations was conducted in the model diatom Phaeodactylum tricornutum. Out of 10 402 predicted genes, 2491 and 405 genes were significantly upregulated or downregulated respectively. Unsurprisingly, genes associated with phosphate uptake were upregulated, such as the phosphate transporters and alkaline phosphatases. Genes encoding stress-shock proteins were accordingly upregulated, including genes associated with stress-responsive proteins, signal transduction and secondary metabolism. Additionally, genes related to protein translation, carbon fixation, glycolysis and the citric acid cycle were also upregulated. Genes associated with gene transcription were downregulated, thereby resulting in the upregulation of translation to compensate for the limited supply of messenger RNA. The downregulation of genes related to ß-oxidation could contribute to the accumulation of fatty acids. Accordingly, triacylglycerols, which are important for energy storage, were determined to increase by 1.65-fold. Intracellular membranes, other than chloroplast membranes, tended to be dispersed; this finding was in accordance with the increased transcription of a total of 11 genes encoding putative phospholipases. Taken together, this work revealed the coordination of multiple metabolic pathways and certain key genes in the adaptation of P. tricornutum to phosphorus stress.

[Antidepressant activities of flavonoids from Glycyrrhiza uralensis and its neurogenesis protective effect in rats].

Adult rats chronic unpredictable stress model of depression (CUS) was adopted to elucidate the antidepressant pharmacological activity and related neurogenesis protective effect of the total flavonoids extract (licorice flavonoids, LF) from the Glycyrrhiza uralensis Fisch. cultivated locally in Ningxia. The rats were exposed to 9 kinds of unpredictable sequence of stressors and were given flavonoids (300 mg x kg(-1), 100 mg x kg(-1) and 30 mg x kg(-1)) for 28 days. The antidepressant effect was elucidated by open field test, forced swimming test and tail suspension test. The level of serum corticosterone was detected by radioimmunoassay. 5'-Bromo-2'-deoxyuridine (BrdU) labeling experiments was employed to study the neurogenesis protective activities. The flavonoids can increase the sum of line crosses and number of rears, and decrease the number of fecal boli produced in the open field test of the CUS rats. Also the flavonoids can decrease the immobility time in forced swim test as well as in the tail suspension test. In addition, the flavonoids (300 mg x kg(-1)) can decrease the serum corticosterone level of the CUS rats, and increase the number of the new born BrdU positive progenitor cells at the subgranular zone (SGZ) of dentate gyrus (DG) region in hippocampus. The results demonstrated that the total flavonoids extract from the cultivated Glycyrrhiza uralensis Fisch. could produce the anti-depressive effect on chronic unpredictable stress of depression model rats and its mechanism may be associated with its neurogenesis protective effect.

Inhibition of the growth of Alexandrium tamarense by algicidal substances in Chinese fir (Cunninghamia lanceolata).

The wood sawdust from Chinese fir (Cunninghamia lanceolata) exhibited stronger inhibition on the growth of Alexandrium tamarense than those from alder (Alnus cremastogyne), pine (Pinus massoniana), birch (Betula alnoides) and sapele (Entandrophragma cylindricum). The water extract, acetone-water extract and essential oil from fir sawdust were all shown to inhibit the growth of A. tamarense. The inhibition of fir essential oil was the strongest among all the above wood sources while the half effective concentration was only 0.65 mg/L. These results suggested that the fir essential oil may play an important role in the algicidal effect of Chinese fir.

[Inhibitory effects and chemical basis of cornstalk on the growth of Alexandrium tamarense].

To provide more information on new algaecide with high efficiency, ecological safety and selectivity, effects of corn stem and leaves on the growth of Alexandrium tamarense were observed. The roles of microorganism in the inhibition were assessed. The inhibitory activities of different solvent extracts from the corn leaves were discussed and the potential antialgal chemicals in corn leaves were analyzed by GC-MS. Cornstalk is shown to have distinctly inhibitory effect on A. tamarense, and the inhibition of corn leaves is stronger than that of the corn stem. 0.5 g/L of the corn leaves inhibit A. tamarense remarkably in cell density of 1.69 x 10(6) cells/L. There are little differences in antialgal action between asepsis leaves and rude leaves, suggesting that some antialgal compounds from leaves may be responsible for the inhibition and that microorganisms from leaves have little effect on the inhibition. The petroleum ether and dichloromethane extracts from corn leaves are shown to have stranger inhibition on A. tamarense than ethyl acetate and n-butanol extracts. CC-MS shows that extracts with high inhibitory activities contain many fatty acids such as linoleic acid, linolenic acid, and palmitic acid, etc. These results suggest that corn leaves have some inhibitory effect on A. tamarense and fatty acids may be responsible for the inhibition.

[Inhibitory effects and chemical basis of Eucalyptus orelliana wood meals on the growth of Alexandrium tamarense].

To provide information on the screen of newly and efficient algaecides in controlling harmful algal blooms (HABs), the effects of wood meals from Eucalyptus torelliana, Eucalyptus urophylla, Eucalyptus exserta on the growth of Alexandrium tamarense were observed and the chemical basis of the antialgal effect was discussed. The results show that the inhibitory activities of the three wood meals are different, and E. torelliana wood meals have the highest inhibitory activity. There are little differences in antialgal action between asepsis and rude wood meals, suggesting that some antialgal compounds from wood meals may be responsible for the inhibition and that microorganisms from wood meals have little effect on the inhibition. The acetone-water extract from E. torelliana wood meals is shown to have stronger inhibition on A. tamarense than that from ethyl acetate, water and methanol extracts. The acetone-water extract from E. torelliana wood meals was further divided into extract A, B, C and D and the inhibitory activities were compared. The extract D is shown to have highest inhibitory activity. 3 mg/L of the extract appears 81% inhibition rate to A. tamarense in the 3rd day. GC-MS show that extract D contains mostly ketones such as 4-hydroxy-3,5,6-trimethyl-4-(3-oxo-1-butenyl)-2-cyclohexen-1-one and 5,6,7,7a-tetrahydro-4,4,7a-trimethyl-2(4H)-benzofuranone. These results suggest that wood meals from E. torelliana had certain inhibitory effect on A. tamarense, and that ketones may be responsible for the inhibition.

[Effects of different phosphorus sources on the growth and toxin production of Prorocentrum lima].

To explore the nutrient properties of Prorocentrum lima and biosynthesis mechanism of diarrhetic shellfish poison (DSP), the growth and activities of alkaline phosphatase of Prorocentrum lima were observed under different phosphorus sources. DSP productions were also analyzed. The maximum growth rate (micro(max)) was slightly lower under beta-sodium glycerophosphate than those under NaH2PO4 and ATP as phosphorus sources, respectively. The maximum biomass (X) under ATP was higher than those under NaH2PO4 and beta-sodium glycerophosphate as the phosphorus sources, respectively. When the concentration of NaH2PO4 was below 2 micromol/L, the activity of alkaline phosphatase increased significantly. However, the activities were much low in the all treatments when beta-sodium glycerophosphate used as phosphorus source, whereas the activities increased with the concentration of ATP when ATP used as phosphorus source. The level of okadaic acid (OA) in Prorocentrum lima at the stationary phase under beta-sodium glycerophosphate was higher than those under NaH2PO4 and ATP. These suggested that beta-sodium glycerophosphate could be utilized directly by Prorocentrum lima with lower efficiency; ATP could induce alkaline phosphatase to produce inorganic phosphate for algae. DSP production in Prorocentrum lima were different under various phosphate sources, beta-sodium glycerophosphate enhanced production of DSP. The difference in DSP production might be related with the physiological state of Prorocentrum lima.

[Hypnotic effect of Chinese materia medica Cynanchum chinese in mice].

OBJECTIVE: To study the central pharmacological effect of the water and chloroform-extract compounds from C. chinese in mice. METHOD: The independent activity test and the hypnotic synergism test by sub-threshold hypnotic dosage of pentobarbital were employed to evaluate the central pharmacological effect of the extract-compounds, and the minimal neurotoxicity was evaluated by the rotorod test. RESULT: the extract-compounds exhibited significant dose-related inhibition effect of the spontaneous motor activity in mice after intraperitoneal administration. And the two extract-compounds promoted the hypnotic effect by sub-threshold hypnotic dosage administration of pentobarbital, and produced ED50 value of 2.36 g kg (-1) and 0.75 g kg(-1), respectively. Also, both extract-compounds showed no neurotoxicity in the experiment. CONCLUSION: The extract compounds from C. chinese showed inhibitional effect on CNS.