In Vitro and In Vivo Anti-Inflammatory Effects of Cannabidiol Isolated from Novel Hemp (Cannabis sativa L.) Cultivar Pink Pepper.

Cannabis sativa L. contains more than 80 cannabinoids, among which cannabidiol (CBD) is the main neuroactive component. We aimed to investigate the anti-inflammatory efficacy of CBD in vitro and in vivo isolated from "Pink pepper", a novel hemp cultivar, by repeating the method of selecting and cultivating individuals with the highest CBD content. We investigated the effects of CBD on inflammatory markers elevated by lipopolysaccharide (LPS) treatment in RAW 264.7 mouse macrophage cells through Western blot and RT-PCR. In addition, we confirmed these effects through the ELISA of inflamed paw tissue of a λ-carrageenan-induced mouse edema model that received an oral administration of CBD. CBD inhibited the LPS-induced phosphorylation of NF-κB and MAPK in RAW 264.7 and exhibited anti-inflammatory effects by participating in these pathways. In our in vivo study, we confirmed that CBD also inhibited the inflammatory mediators of proteins extracted from edematous mouse paw tissue. These results show that CBD isolated from "Pink pepper" exhibits potent anti-inflammatory effects. These anti-inflammatory effects of CBD have pharmacological and physiological significance, highlighting the industrial value of this novel cultivar.

Conversion Characteristics of Some Major Cannabinoids from Hemp (Cannabis sativa L.) Raw Materials by New Rapid Simultaneous Analysis Method.

This study was carried out to develop a high-performance liquid chromatography method for short-time analysis of the main cannabinoids in the inflorescence of hemp (Cannabis sativa L.). We also performed decarboxylation of the raw material using our advanced analysis technique. In this study, the UV spectrum was considered to analyze each of the four common cannabinoids, solvents, and samples, where the uniform elution of acidic cannabinoids without peak tailing and acids was tested. Optimal results were obtained when readings were taken at a wavelength of 220 nm using water and methanol containing trifluoroacetic acid as mobile phases in a solvent gradient system. The established conditions were further validated by system suitability, linearity, precision, detection limit, and quantitation limit tests. The decarboxylation index (DT50) confirmed that Δ9-THCA decarboxylated faster than CBDA, and both maintained a linear relationship with time and temperature. In addition, the loss of cannabidiol was better prevented during the decarboxylation process in the natural state than in the extracted state.

Effect of Light Quality on Seed Potato (Solanum tuberose L.) Tuberization When Aeroponically Grown in a Controlled Greenhouse.

A plant factory equipped with artificial lights is a comparatively new concept when growing seed potatoes (Solanum tuberosum L.) for minituber production. The shortage of disease-free potato seed tubers is a key challenge to producing quality potatoes. Quality seed tuber production all year round in a controlled environment under an artificial light condition was the main purpose of this study. The present study was conducted in a plant factory to investigate the effects of distinct spectrum compositions of LEDs on potato tuberization when grown in an aeroponic system. The study was equipped with eight LED light combinations: L1 = red: blue: green (70 + 25 + 5), L2 = red: blue: green (70 + 20 + 10), L3 = red: blue: green (70 + 15 + 15), L4 = red: blue: green (70 + 10 + 20), L5 = red: blue: far-red (70 + 25 + 5), L6 = red: blue: far-red (70 + 20 + 10), L7 = red: blue: far-red (70 + 15 + 15), L8 = red: blue: far-red (70 + 10 + 20), and L9 = natural light with 300 µmol m-2 s-1 of irradiance, 16/8 h day/night, 65% relative humidity, while natural light was used as the control treatment. According to the findings, treatment L4 recorded a higher tuber number (31/plant), tuber size (>3 g); (9.26 ± 3.01), and GA3 content, along with better plant growth characteristics. Moreover, treatment L4 recorded a significantly increased trend in the stem diameter (11.08 ± 0.25), leaf number (25.32 ± 1.2), leaf width (19 ± 0.81), root length (49 ± 2.1), and stolon length (49.62 ± 2.05) compared to the control (L9). However, the L9 treatment showed the best performance in plant fresh weight (67.16 ± 4.06 g) and plant dry weight (4.46 ± 0.08 g). In addition, photosynthetic pigments (Chl a) (0.096 ± 0.00 mg g-1, 0.093 ± 0.00 mg g-1) were found to be the highest in the L1 and L2 treatments, respectively. However, Chl b and TCL recorded the best results in treatment L4. Finally, with consideration of the plant growth and tuber yield performance, treatment L4 was found to have the best spectral composition to grow quality seed potato tubers.

In Vitro and In Vivo Anti-Inflammatory Potential of Cannabichromene Isolated from Hemp.

Cannabichromene (CBC), a non-psychoactive cannabinoid found in Cannabis sativa, has recently been shown to possess several medicinal properties. However, how CBC produces anti-inflammatory effects and the mechanisms of this remain poorly studied. Therefore, we extracted and purified the CBC from the Cannabis sativa cv. pink pepper (hemp cultivar). The efficacy of CBC in reducing inflammation in RAW 264.7 macrophages and a λ-carrageenan-induced mouse model was then evaluated. CBC had no cytotoxicity up to a concentration of 20 µM and inhibited nitric oxide production by approximately 50% at a concentration of 20 µM. In addition, CBC treatment significantly inhibited causes of inflammation such as inducible nitric oxide synthase (iNOS), interleukin-1ß (IL-1ß), IL-6, and tumor necrosis factor-α (TNF-α) at both the mRNA and protein levels. Moreover, CBC suppressed LPS-stimulated inflammation in RAW 264.7 cells by downregulating the nuclear factor kappa B (NF-kB) and mitogen-activated protein kinase pathways (MAPK). Furthermore, our in vivo experiments confirmed that the λ-carrageenan-induced increase in the levels of the cytokines iNOS, IL-1ß, and IL-6 was abrogated following treatment with CBC. Therefore, CBC has potential anti-inflammatory effects and may be useful for preventing or treating inflammation.

Exogenous putrescine attenuates the negative impact of drought stress by modulating physio-biochemical traits and gene expression in sugar beet (Beta vulgaris L.).

Drought tolerance is a complex trait controlled by many metabolic pathways and genes and identifying a solution to increase the resilience of plants to drought stress is one of the grand challenges in plant biology. This study provided compelling evidence of increased drought stress tolerance in two sugar beet genotypes when treated with exogenous putrescine (Put) at the seedling stage. Morpho-physiological and biochemical traits and gene expression were assessed in thirty-day-old sugar beet seedlings subjected to drought stress with or without Put (0.3, 0.6, and 0.9 mM) application. Sugar beet plants exposed to drought stress exhibited a significant decline in growth and development as evidenced by root and shoot growth characteristics, photosynthetic pigments, antioxidant enzyme activities, and gene expression. Drought stress resulted in a sharp increase in hydrogen peroxide (H2O2) (89.4 and 118% in SBT-010 and BSRI Sugar beet 2, respectively) and malondialdehyde (MDA) (35.6 and 27.1% in SBT-010 and BSRI Sugar beet 2, respectively). These changes were strongly linked to growth retardation as evidenced by principal component analysis (PCA) and heatmap clustering. Importantly, Put-sprayed plants suffered from less oxidative stress as indicated by lower H2O2 and MDA accumulation. They better regulated the physiological processes supporting growth, dry matter accumulation, photosynthetic pigmentation and gas exchange, relative water content; modulated biochemical changes including proline, total soluble carbohydrate, total soluble sugar, and ascorbic acid; and enhanced the activities of antioxidant enzymes and gene expression. PCA results strongly suggested that Put conferred drought tolerance mostly by enhancing antioxidant enzymes activities that regulated homeostasis of reactive oxygen species. These findings collectively provide an important illustration of the use of Put in modulating drought tolerance in sugar beet plants.

Cannabinoid accumulation in hemp depends on ROS generation and interlinked with morpho-physiological acclimation and plasticity under indoor LED environment.

Manipulation of growth and development of cannabis (Cannabis sativa L.) has received considerable interest by the scientific community due to its high value in medicinal and recreational use worldwide. This study was conducted to investigate the effects of LED spectral changes on reactive oxygen species (ROS) and cannabinoid accumulation by provoking growth, pigmentation, photosynthesis, and secondary metabolites production of cannabis grown in an indoor environment. After three weeks of vegetative growth under greenhouse condition, plants were further grown for 90 days in a plant factory treated with 4 LED light compositions with a canopy-level photosynthetic photon flux density (PPFD) of 300 µmol m-2 s-1 for 16 h. Photosynthetic pigments and photosynthetic rate were linearly increased up to 60 days and then sharply decreased which was found most prominent in L3: MB 240 (Red 85% + Blue 15%) and L4: PF 240 (Red 70% + Blue 30%) LED light compositions. A high concentration of H2O2 was also observed in L3 and L4 treatments which provoked lipid peroxidation in later growth stage. In addition, higher accumulation of cannabinoid was observed under L4 treatment in most cases. It is also evident that higher ROS created a cellular stress in plant as indicated by higher osmolyte synthesis and enzyme activity which initiate quick maturation along with higher cannabinoids accumulation in cannabis plant. Therefore, it can be concluded that ROS metabolism has a crucial role in morpho-physiological acclimation and cannabinoid accumulation in hemp plants. The findings of this study provide further insight on the use of LED light to maximize the production of cannabinoid.

Exogenous Putrescine Enhances Salt Tolerance and Ginsenosides Content in Korean Ginseng (Panax ginseng Meyer) Sprouts.

The effect of exogenously applied putrescine (Put) on salt stress tolerance was investigated in Panax ginseng. Thirty-day-old ginseng sprouts were grown in salinized nutrient solution (150 mM NaCl) for five days, while the control sprouts were grown in nutrients solution. Putrescine (0.3, 0.6, and 0.9 mM) was sprayed on the plants once at the onset of salinity treatment, whereas control plants were sprayed with water only. Ginseng seedlings tested under salinity exhibited reduced plant growth and biomass production, which was directly interlinked with reduced chlorophyll and chlorophyll fluorescence due to higher reactive oxygen species (hydrogen peroxide; H2O2) and lipid peroxidation (malondialdehyde; MDA) production. Application of Put enhanced accumulation of proline, total soluble carbohydrate, total soluble sugar and total soluble protein. At the same time, activities of antioxidant enzymes like superoxide dismutase, catalase, ascorbate peroxidase, guaiacol peroxidase in leaves, stems, and roots of ginseng seedlings were increased. Such modulation of physio-biochemical processes reduced the level of H2O2 and MDA, which indicates a successful adaptation of ginseng seedlings to salinity stress. Moreover, protopanaxadiol (PPD) ginsenosides enhanced by both salinity stress and exogenous Put treatment. On the other hand, protopanaxatriol (PPT) ginsenosides enhanced in roots and reduced in leaves and stems under salinity stress condition. In contrast, they enhanced by exogenous Put application in all parts of the plants for most cases, also evidenced by principal component analysis. Collectively, our findings provide an important prospect for the use of Put in modulating salinity tolerance and ginsenosides content in ginseng sprouts.

Cannabinoids Accumulation in Hemp (Cannabis sativa L.) Plants under LED Light Spectra and Their Discrete Role as a Stress Marker.

Hemp adaptability through physiological and biochemical changes was studied under 10 LED light spectra and natural light in a controlled aeroponic system. Light treatments were imposed on 25 days aged seedlings for 16 h daily (300 µmol m-2 s-1) for 20 days. Plant accumulated highest Cannabidiol (CBD) in R7:B2:G1 light treatment, with relatively higher photosynthetic rate and lower reactive oxygen species, total phenol content, total flavonoid content, DPPH radical scavenging capacity, and antioxidant enzymatic activities. Tetrahydrocannabinol (THC) also accumulated at a higher level in white, R8:B2, and R7:B2:G1 light with less evidence of stress-modulated substances. These results indicated that CBD and THC have no or little relation with light-mediated abiotic stress in hemp plants. On the contrary, Tetrahydrocannabinolic acid (THCA) was accumulated higher in R6:B2:G1:FR1 and R5:B2:W2:FR1 light treatment along with lower photosynthetic rate and higher reactive oxygen species, total phenol content, total flavonoid content, DPPH radical scavenging capacity, and antioxidant enzymatic activities. However, Cannabidiolic acid (CBDA) was accumulated higher in R6:B2:G1:FR1 light treatment with higher stress-modulated substances and lower physiological traits. CBDA was also accumulated higher in R8:B2 and R7:B2:G1 light treatments with less evidence of stress-modulated substances. Besides, Greenlight influenced CBD and CBDA synthesis where FR and UV-A (along with green) play a positive and negative role in this process. Overall, the results indicated that the treatment R7:B2:G1 enhanced the medicinal cannabinoids most, and the role of THCA as a stress marker is more decisive in the hemp plant than in other cannabinoids under attributed light-mediated stress.

Production of Potato (Solanum tuberosum L.) Seed Tuber under Artificial LED Light Irradiation in Plant Factory.

Plant production in a plant factory is an innovative and smart idea to grow food anytime, anywhere, regardless of the outer environment. However, potato pre-basic seed tuber (PBST) production in a plant factory is a comparatively new initiative. Therefore, the aim of this study was to optimize the artificial LED light spectrum to produce PBST in a plant factory. Two potato varieties such as Golden king (V48) and Chungang (V41) were grown in soil substrate under different combination of artificial LED light combinations (such as red+blue+far-red, red+blue+white, blue+far-red, blue+white, red+far-red, and red+white) maintaining photosynthetic photon flux density (PPFD) of 100 mol m-2s-1, temperature 23/15 °C (day/night), and relative humidity 70%. The study revealed that, overall, potato plant growth (viz.; plant height, node number, leaf number, leaf length and width, fresh and dry weight) was enhanced by the red+far red light for both potato varieties. The total seed tuber number per plant was higher in red+blue+white light for V48, and red+far-red for V41. The fresh tuber weight was the highest in the red+blue+far-red light for V48 and red+blue+white for V41. The highest accumulated photosynthetic pigment (total Chlorophyll, Chlorophyll a, b and Carotenoid) was observed in red+blue+white light for both varieties. The total carbohydrate content and total sucrose content were higher in red+blue+far red and red +far red light treatment for V48 and V41, respectively. Finally, considering all factors, it is concluded that the red+blue+white light combination is deemed to be appropriate for the potato PBST production in plant factory conditions.

Physiological and Biochemical Changes in Sugar Beet Seedlings to Confer Stress Adaptability under Drought Condition.

The present study was conducted to examine the adaptability of 11 sugar beet cultivars grown under drought stress in the controlled glasshouse. The treatment was initiated on 30-day-old sugar beet plants where drought stress was made withholding water supply for consecutive 10 days while control was done with providing water as per requirement. It was observed that drought stress expressively reduced plant growth, photosynthetic pigments, and photosynthetic quantum yield in all the cultivars but comparative better results were observed in S1 (MAXIMELLA), S2 (HELENIKA), S6 (RECODDINA), S8 (SV2347), and S11 (BSRI Sugarbeet 2) cultivars. Besides, osmolytes like proline, glycine betaine, total soluble carbohydrate, total soluble sugar, total polyphenol, total flavonoid, and DPPH free radical scavenging activity were remarkably increased under drought condition in MAXIMELLA, HELENIKA, TERRANOVA, GREGOIA, SV2348, and BSRI Sugar beet 2 cultivars. In contrast, activities of enzymes like superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) were significantly decreased in all, while the cultivars SV2347, BSRI Sugar beet 1 and BSRI Sugar beet 2 were found with increased ascorbate peroxidase (APX) activity under drought condition. In parallel, polyphenol oxidase (PPO) was increased in all cultivars except HELENIKA. Overall, the cultivars HELENIKA, RECODDINA, GREGOIA, SV2347, SV2348, BSRI Sugar beet 1, and BSRI Sugar beet 2 were found best fitted to the given drought condition. These findings would help further for the improvement of stress adaptive sugar beet cultivars development in the breeding program for drought-prone regions.

Comparative Study of Piper sylvaticum Roxb. Leaves and Stems for Anxiolytic and Antioxidant Properties Through in vivo, in vitro, and in silico Approaches.

Piper sylvaticum Roxb. is traditionally used by the indigenous people of tropical and subtropical countries like Bangladesh, India, and China for relieving the common cold or a variety of chronic diseases, such as asthma, chronic coughing, piles, rheumatic pain, headaches, wounds, tuberculosis, indigestion, and dyspepsia. This study tested anxiolytic and antioxidant activities by in vivo, in vitro, and in silico experiments for the metabolites extracted (methanol) from the leaves and stems of P. sylvaticum (MEPSL and MEPSS). During the anxiolytic evaluation analyzed by elevated plus maze and hole board tests, MEPSL and MEPSS (200 and 400 mg/kg, body weight) exhibited a significant and dose-dependent reduction of anxiety-like behavior in mice. Similarly, mice treated with MEPSL and MEPSS demonstrated dose-dependent increases in locomotion and CNS simulative effects in open field test. In addition, both extracts (MEPSL and MEPSS) also showed moderate antioxidant activities in DPPH scavenging and ferric reducing power assays compared to the standard, ascorbic acid. In parallel, previously isolated bioactive compounds from this plant were documented and subjected to a molecular docking study to correlate them with the pharmacological outcomes. The selected four major phytocompounds displayed favorable binding affinities to potassium channel and xanthine oxidoreductase enzyme targets in molecular docking experiments. Overall, P. sylvaticum is bioactive, as is evident through experimental and computational analysis. Further experiments are necessary to evaluate purified novel compounds for the clinical evaluation.

Unveiling Pharmacological Responses and Potential Targets Insights of Identified Bioactive Constituents of Cuscuta reflexa Roxb. Leaves through In Vivo and In Silico Approaches.

Cuscuta reflexa Roxb. is traditionally used by the indigenous communities of Bangladesh to treat different diseases, such as pain, edema, tumor, jaundice, and skin infections. This study tested neuro-pharmacological, anti-nociceptive, and antidiarrheal activities by in vivo and in silico experiments for the metabolites extracted (methanol) from the leaves of Cuscuta reflexa (MECR). During the anxiolytic evaluation analyzed by elevated plus maze and hole board tests, MECR (200 and 400 mg/kg) exhibited a significant dose-dependent reduction of anxiety-like behavior in mice. Similarly, mice treated with MECR demonstrated a dose-dependent decrease in the time of immobility in both forced swimming and tail suspension tests. In addition, anti-nociceptive activity was assessed by the chemical-induced (acetic acid and formalin) pain models. In both cases, 400 mg/kg was found to be most effective and significantly (p < 0.001) inhibited acetic acid stimulated writhing and formalin-induced licking (pain response) in mice. Furthermore, antidiarrheal efficacy determined by the castor-oil induced diarrheal model manifested an evident inhibition of diarrheal stool frequency. In parallel, previously isolated bioactive compounds were documented based on the biological activities and subjected to in silico studies to correlate with the current pharmacological outcomes. The selected isolated compounds (15) displayed favorable binding affinities to potassium channels, human serotonin receptor, COX-1, COX-2, M3 muscarinic acetylcholine receptor, and 5-HT3 receptor proteins. Additionally, the ADME/T and toxicological properties were justified to unveil their drug-like properties and toxicity level. Overall, Cuscuta reflexa is bioactive and could be a potential source for the development of alternative medicine.

Intervention in Neuropsychiatric Disorders by Suppressing Inflammatory and Oxidative Stress Signal and Exploration of In Silico Studies for Potential Lead Compounds from Holigarna caustica (Dennst.) Oken leaves.

Holigarna caustica (Dennst.), a popular plant used in folk medicine in Bangladesh, is often used by the local folk practitioner to treat a variety of chronic diseases. The present research is an attempt to find out an innovative therapeutic prospect for the management of neuropsychiatric disorders. The methanol extract of H. caustica leaves (MEHC) were utilized on various behavioral tests for assessing anxiolytic, anti-depressant, and anti-inflammatory activities. The antioxidant potentials and quantitative phytochemicals were evaluated through spectrophotometric methods. Results revealed that treatment of MEHC (200 and 400 mg/kg) significantly reduced anxiety like behaviors in mice, particularly, 400 mg/kg efficiently improved % of entries and time spent (p < 0.05) in the open arms in elevated plus maze test, whereas, superior head dipping tendency (p < 0.05) was observed in hole-board test. In contrast, mice treated with 200 mg/kg revealed better anxiolytic effect in both open field and hole-cross tests. During antidepressant evaluation, mice administrated with MEHC exhibited active behaviors (swimming and struggling) in forced swimming and tail suspension tests. In parallel, MEHC manifested a noteworthy (p < 0.001) suppression of inflammatory response induced by histamine. The MEHC also showed strong antioxidant activities in 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) (IC50: 57.64 µg/mL) scavenging, H2O2 (IC50: 51.60 µg/mL) scavenging, and ferric reducing power assay. The levels of total phenol, flavonoid, flavonol, condensed tannin, and antioxidant were estimated as higher in MEHC. Moreover, 11 compounds were documented as bioactive, displayed good binding affinities to potassium channel receptor, human serotonin receptor, cyclooxygenase (COX-1 and 2), and xanthine oxidoreductase enzyme targets in molecular docking experiments. Furthermore, ADME/T and Prediction of Activity Spectra for Substances (PASS) analyses exposed their drug-likeness, nontoxic upon consumption, and likely pharmacological actions. Overall, the H. caustica is potentially bioactive as evident by in vivo, in vitro, and computational analysis. Our findings support the folkloric value of this plant, which may provide a potential source towards developing drug leads.

Clinical Characteristics and Treatment Outcome of Peginterferon Plus Ribavirin in Patients Infected with Genotype 6 Hepatitis C Virus in Korea: A Multicenter Study.

BACKGROUND/AIMS: Because of the limited geographic distribution, there have been insufficient data regarding hepatitis C virus (HCV) genotype 6 in Korea. This study aimed to investigate the clinical characteristics and available treatment outcomes of patients with genotype 6 HCV in Korea. METHODS: From 2004 to 2014, data were collected from Korean patients infected with genotype 6 HCV in eight hospitals. RESULTS: Thirty-two patients had genotype 6 HCV. The median age was 44 years, and 6c was the most common subtype. The baseline median alanine transaminase level was 88 (21 to 1,019) IU/mL, and the HCV RNA level was 1,405,000 (96,500 to 28,844,529) IU/mL. Twenty-five patients were treated with peginterferon (PEG-IFN) and ribavirin. Three follow-up losses occurred. Additionally, 13 patients attained a sustained virologic response (SVR), seven patients relapsed, and two patients exhibited a null response. The SVR rates were 40% and 75% for the 24- and more than 48-week treatments, respectively, and five of the six patients who achieved a rapid virologic response (RVR) attained a SVR. CONCLUSIONS: Korean patients infected with genotype 6 HCV are relatively young, and 6c is the most common subtype. When treated with PEG-IFN and ribavirin, the SVR rate was 52%. Similar to other genotypes, a longer duration of treatment and attainment of RVR are important for SVR.

Prediction of treatment outcome in chronic hepatitis C patients based on early viral dynamics during high-dose induction interferon and ribavirin therapy.

OBJECTIVE: In chronic hepatitis C, early viral load decline after interferon administration is dose dependent and reflects the intrinsic viral susceptibility to the antiviral action of interferon. We examined whether the augmented suppression of susceptible viral loads by high-dose induction interferon could possibly discriminate responsive patients from non-responsive patients at an early stage of treatment. METHODS: Fifty-nine chronic hepatitis C patients were randomly allocated to receive one of two treatment regimens; 3 MU interferon three times weekly plus ribavirin 1,000 mg/day for 24 weeks in the CR group (n = 30), and the same regimen as in the CR group except 10 MU interferon daily for the first week in the HR group (n = 29). Changes in viral loads during the first week of treatment were analyzed in terms of sustained virological response (SVR). RESULTS: The positive predictive values of undetectable (<100 IU/ml) or low serum HCV RNA (<2,000 IU/ml) after 1 week of treatment for SVR were 100% in both treatment groups, whereas the negative predictive values of the low viral titer were 91% in the HR group and 70% in the CR group. CONCLUSION: One-week virological response to high-dose induction interferon/ribavirin combination therapy is more predictive of SVR than conventional combination therapy in chronic hepatitis C.