Kultur suspensi sel tanaman gajah beranak (Goniothalamus tapis Miq) terhadap kandungan zat goniotalamin

Zat goniotalamin pada tanaman gajah beranak (Goniothalamus tapis) merupakan obat alternatif penyembuhan kanker. Penelitian bertujuan untuk mendapatkan zat goniotalamin melalui kultur kalus dan kultur suspensi sel. Metode penelitian eksperimen Rancangan Acak Lengkap (RAL) dengan kombinasi 2,4-D (1-10 mgL-1) dan BAP (0,5-2 mgL-1) menggunakan eksplan batang muda, terdiri dari 17 perlakuan dan 3 kali ulangan. Analisis data menggunakan Analysis of Variances dan uji lanjut Duncan Multiple Range Test (DMRT) taraf 5%. Hasil menunjukkan bahwa kultur kalus G. tapis pada media 5,0 mg L-1 2,4-D + 1 mg L-1 BAP adalah yang terbaik dengan waktu muncul kalus 28,33 hari dan persentase pembentukan kalus 100%. Kalus untuk kultur suspensi sel bertekstur remah dan berwarna kuning kehijauan. Kultur suspensi sel menghasilkan pertumbuhan sel yang cepat, tidak lembek berair dan mudah dipisahkan. Hasil kualitatif Kromatografi Lapis Tipis kultur suspensi sel sangat jelas, bersih dan terdapat potensi kandungan zat goniotalamin pada perlakuan 2,4-D 5 mg L-1 + BAP 0,5 mg L-1, 2,4-D 5 mg L-1 + BAP 1 mg L-1, 2,4-D 5 mg L-1 + BAP 2 mg L-1, 2,4-D 10 mg L-1 + BAP 0,5 mg L-1 dan 2,4-D 10 mg L-1 + BAP 1 mg L-1. Hasil kuantitatif zat goniotalamin dengan Kromatografi Cair Prestasi Tinggi terdapat pada perlakuan 2,4-D 5,0 mgL-1 + BAP 1 mg L-1 yaitu 9,57 mg g-1.The goniothalamine compound on Goniothalamus tapis is an alternative cancer medicine. This study aimed to obtain gonotalamin through callus culture and suspension cell culture. The experiment research method was Completely Randomized Design (CRD) with a combination of 2.4-D (1-10 mg L-1) and BAP (0.5-2 mg L-1) using young stem explants consisting of 17 treatments with 3 replications. Data analysis used ANOVA and DMRT at 5%. The results showed that G. tapis callus culture on 5.0 mg L-1 2.4-D + 1 mg L-1 BAP was the best treatment medium with callus emergence time of 28.33 days and percentage of callus formation 100%. The callus used for suspension cell culture was friable and greenish-yellow in color. Suspension cell culture resulted in rapid cell growth, was not fleshy, and easily separated. The quality test by Thin Layer Chromatography (TLC) from suspension cell culture resulted very clear, clean, and potential content of goniothalamin found in treatments 2.4-D 5.0 mg L-1 + BAP 0.5 mg L-1, 2.4-D 5.0 mg L-1 + BAP 1 mg L-1, 2.4-D 5.0 mg L-1 + BAP 2 mg L-1, 2.4-D 10 mg L-1 + BAP 0.5 mg-1 and 2.4-D 10 mg-1 + BAP 1 mg-1. The quantitative results of the best goniotalamine compounds in cell suspension cultures using High-Performance Liquid Chromatography (HPLC) on medium 2,4-D 5.0 mgL-1 + BAP 1 mg L-1 ie 9.57 g-1.

Rosmarinic acid production in cell suspension cultures of Ehretia asperula Zollinger & Moritz

Plant cell cultures provide an alternative means for producing secondary compounds in food, cosmetic and pharmaceutical industries. Ehretia asperula Zollinger & Moritzi is used as a traditional medicine for the treatment of liver detoxification, ulcers, tumors, inflammation and enhancing the body's resistance in Vietnam. The study was carried out to select suitable callus line for cell suspension cultures of E. asperula Zollinger & Moritzi and investigate the effects of inoculum size, rotation speed and naphthalene acetic acid (NAA) on the proliferation of cell suspension cultures. In addition, the influence of light intensity on the growth and rosmarinic acid (RA) biosynthesis of cell suspension was also surveyed. After 4 weeks of culture, the white to pale yellow friable callus expanded significantly with a fresh weight (FW) of 0.788 g and a high RA content of 2.062 mg/g FW. An appropriate medium for cell proliferation was the liquid B5 medium, which contained 30 g/l glucose, 0.1 mg/l benzyl adenine (BA) and 0.4 mg/l NAA. The results also demonstrated that a 1:20 ratio (w/v) inoculum size, darkness and rotation speed of 90 rpm were the optimal conditions for the proliferation and RA accumulation to 188.217 mg/l in 4 weeks of culture. These findings showed that E. asperula Zollinger & Moritzi cell suspension cultures could be a potential alternative approach for RA production in vitro.

Producing Medicinal Phytochemicals from Phyllanthus acuminatus in Plant - Cell Suspension Cultures

Pharmaceutical use is not feasible for important medicinal compounds derived from certain plant materials, including Phyllanthus acuminatus roots, due to their low natural abundance. New technologies in non-traditional biomass generation are needed to produce these remarkable natural compounds. Therefore, this article describes a methodology for establishing Phyllanthus acuminatus plant-cell suspensions from callus cultures: An evaluation on inoculum concentration and agitation speed displayed significant changes in plant cell growth kinetics. It was determined that treatment with 2 g of inoculum in 25 mL of medium and 100 rpm agitation creates the best conditions for generating thick cell suspensions. Likewise, treatment with 2 g of inoculum and 120 rpm agitation produces the best conditions for establishing fine cell suspensions. Phytochemical comparison through high-resolution mass spectrometry of P. acuminatus roots and plant cell suspension extracts confirmed presence in the plant cell culture of multiple phyllantostatins of pharmaceutical interest. Here, we demonstrate that Phyllanthus acuminatus can be cultured in plant cell suspensions to produce secondary metabolites of medical interest – technology that could be scaled up for implementation in industrial bioprocesses.

LEDitSHAKE: a lighting system to optimize the secondary metabolite content of plant cell suspension cultures

Plant secondary metabolites are widely used in the food, cosmetic and pharmaceutical industries. They can be extracted from sterile grown plant cell suspension cultures, but yields and quality strongly depend on the cultivation environment, including optimal illumination. Current shaking incubators do not allow different light wavelengths, intensities and photoperiods to be tested in parallel. We therefore developed LEDitSHAKE, a system for multiplexed customized illumination within a single shaking incubator. We used 3D printing to integrate light-emitting diode assemblies into flask housings, allowing 12 different lighting conditions (spectrum, intensity and photoperiod) to be tested simultaneously. We did a proof of principle of LEDitSHAKE using the system to optimize anthocyanin production in grapevine cell suspension cultures. The effect of 24 different light compositions on the total anthocyanin content of grapevine cell suspension cultures was determined using a Design of Experiments approach. We predicted the optimal lighting conditions for the upregulation and downregulation of 30 anthocyanins and found that short-wavelength light (blue, UV) maximized the concentration of most anthocyanins, whereas long-wavelength light (red) had the opposite effect. Therefore our results demonstrate proof of principle that the LEDitSHAKE system is suitable for the optimization of processes based on plant cell suspension cultures.

Transcriptomic Changes in Internode Explants of Stinging Nettle during Callogenesis

Callogenesis, the process during which explants derived from differentiated plant tissues are subjected to a trans-differentiation step characterized by the proliferation of a mass of cells, is fundamental to indirect organogenesis and the establishment of cell suspension cultures. Therefore, understanding how callogenesis takes place is helpful to plant tissue culture, as well as to plant biotechnology and bioprocess engineering. The common herbaceous plant stinging nettle (Urtica dioica L.) is a species producing cellulosic fibres (the bast fibres) and a whole array of phytochemicals for pharmacological, nutraceutical and cosmeceutical use. Thus, it is of interest as a potential multi-purpose plant. In this study, callogenesis in internode explants of a nettle fibre clone (clone 13) was studied using RNA-Seq to understand which gene ontologies predominate at different time points. Callogenesis was induced with the plant growth regulators α-napthaleneacetic acid (NAA) and 6-benzyl aminopurine (BAP) after having determined their optimal concentrations. The process was studied over a period of 34 days, a time point at which a well-visible callus mass developed on the explants. The bioinformatic analysis of the transcriptomic dataset revealed specific gene ontologies characterizing each of the four time points investigated (0, 1, 10 and 34 days). The results show that, while the advanced stage of callogenesis is characterized by the iron deficiency response triggered by the high levels of reactive oxygen species accumulated by the proliferating cell mass, the intermediate and early phases are dominated by ontologies related to the immune response and cell wall loosening, respectively.

Phytochemical, Pharmacological, and Biotechnological Study of Ageratina pichinchensis: A Native Species of Mexico

Ageratina pichinchensis (Asteraceae) has been used for a long time in traditional Mexican medicine for treating different skin conditions and injuries. This review aimed to provide an up-to-date view regarding the traditional uses, chemical composition, and pharmacological properties (in vitro, in vivo, and clinical trials) that have been achieved using crude extracts, fractions, or pure compounds. Moreover, for a critical evaluation of the published literature, key databases (Pubmed, Science Direct, and SciFinder, among others) were systematically searched using keywords to retrieve relevant publications on this plant. Studies that reported on crude extracts, fractions, or isolated pure compounds of A. pichinchensis have found a varied range of biological effects, including antibacterial, curative, antiulcer, antifungal, and anti-inflammatory activities. Phytochemical analyses of different parts of A. pichinchensis revealed 47 compounds belonging to chromenes, furans, glycosylated flavonoids, terpenoids, and essential oils. Furthermore, biotechnological studies of A. pichinchensis such as callus and cell suspension cultures have provided information for future research perspectives to improve the production of valuable bioactive compounds.

Knockdown expression of a MYB-related transcription factor gene, OsMYBS2, enhances production of recombinant proteins in rice suspension cells

Background Transgenic plant suspension cells show economic potential for the production of valuable bioproducts. The sugar starvation-inducible rice αAmy3 promoter, together with its signal peptide, is widely applied to produce recombinant proteins in rice suspension cells. The OsMYBS2 transcription factor was shown recently to reduce activation of the αAmy3 promoter by competing for the binding site of the TA box of the αAmy3 promoter with the potent OsMYBS1 activator. In this study, rice suspension cells were genetically engineered to silence OsMYBS2 to enhance the production of recombinant proteins. Results The mouse granulocyte–macrophage colony-stimulating factor (mGM-CSF) gene was controlled by the αAmy3 promoter and expressed in OsMYBS2-silenced transgenic rice suspension cells. Transcript levels of the endogenous αAmy3 and the transgene mGM-CSF were increased in the OsMYBS2-silenced suspension cells. The highest yield of recombinant mGM-CSF protein attained in the OsMYBS2-silenced transgenic suspension cells was 69.8 µg/mL, which is 2.5-fold that of non-silenced control cells. The yield of recombinant mGM-CSF was further increased to 118.8 µg/mL in cultured cells derived from homozygous F5 seeds, which was 5.1 times higher than that of the control suspension cell line. Conclusions Our results demonstrate that knockdown of the transcription factor gene OsMYBS2 increased the activity of the αAmy3 promoter and improved the yield of recombinant proteins secreted in rice cell suspension cultures.