Teucrium polium: Potential Drug Source for Type 2 Diabetes Mellitus

The prevalence of type 2 diabetes mellitus is rising globally and this disease is proposed to be the next pandemic after COVID-19. Although the cause of type 2 diabetes mellitus is unknown, it is believed to involve a complex array of genetic defects that affect metabolic pathways which eventually lead to hyperglycaemia. This hyperglycaemia arises from an inability of the insulin-sensitive cells to sufficiently respond to the secreted insulin, which eventually results in the inadequate secretion of insulin from pancreatic β-cells. Several treatments, utilising a variety of mechanisms, are available for type 2 diabetes mellitus. However, more medications are needed to assist with the optimal management of the different stages of the disease in patients of varying ages with the diverse combinations of other medications co-administered. Throughout modern history, some lead constituents from ancient medicinal plants have been investigated extensively and helped in developing synthetic antidiabetic drugs, such as metformin. Teucrium polium L. (Tp) is a herb that has a folk reputation for its antidiabetic potential. Previous studies indicate that Tp extracts significantly decrease blood glucose levels r and induce insulin secretion from pancreatic β-cells in vitro. Nonetheless, the constituent/s responsible for this action have not yet been elucidated. The effects appear to be, at least in part, attributable to the presence of selected flavonoids (apigenin, quercetin, and rutin). This review aims to examine the reported glucose-lowering effect of the herb, with a keen focus on insulin secretion, specifically related to type 2 diabetes mellitus. An analysis of the contribution of the key constituent flavonoids of Tp extracts will also be discussed

Novel nutritional interventions in the manipulation of energy expenditure and blood glucose regulation in humans

The purpose of the studies described in this thesis was to study the purported metabolic effects of two dietary agents, L-Histidine and Epigallocatechingallate (EGCG). Supplementation of both LHistidine and pure EGCG in animal models has previously shown effects on components of energy balance and blood glucose regulation. Studies of their supplementation among humans are none (in case of L-Histidine) or very limited (in case of EGCG supplementation). The studies presented herein are novel as there have been no previous studies conducted to measure their effects on thermogenesis in humans and the present studies are the first to do. Three studies were performed to investigate metabolic effects of L-Histidine and two studies were performed to investigate metabolic effects of EGCG among healthy men. Duration of supplementation and varying doses were considered in different studies which led to the variation in study designs and protocols. Effects of L-Histidine: The metabolic effects of the essential amino acid L-Histidine (25 mg/kg) on healthy and overweight men were investigated in a series of studies. A pilot study was conducted to develop a protocol for measurement of resting metabolic rate (RMR), mean skin (MST), core temperature (CT) simultaneously. The protocol implemented was successful and was applied in all further studies. This study also examined the effect of L-Histidine supplementation (25 mg/kg) on RMR, MST, CT over a short period (3.25 hours). No conclusive results were obtained from this study. A single blind, placebo controlled cross-over study was then conducted to examine the effects of 10-day supplementation of L-Histidine on RMR, MST, CT and fasting blood glucose (FBG) in healthy male subjects (n = 9). Following L-Histidine ingestion, mean body weight (p = 0.008) and FBG (p = 0.04) were significantly reduced compared to baseline measurements. A randomised, parallel, single blinded and placebo controlled study was conducted following on from this to examine the effects of long-term (8 week, 25 mg/kg/day) intake of L-Histidine on RMR, MST, CT, lipid profile, FBG, insulin, leptin, non-esterified fatty acid (NEFA) and body fat composition in overweight and obese but otherwise healthy men (n = 18 (9 participants each in placebo and L-Histidine group)). Results of this study showed significantly decreased weight after 8 weeks (p = 0.007) compared to baseline, irrespective of the type of supplementation. Oral supplementation of L-Histidine improved FBG concentrations and reduced body weight after 10 days’ ingestion, however, no effects of long-term supplementation was established. Effects of EGCG: The second part of this thesis presents studies performed to investigate metabolic effects of pure EGCG. The first study was randomised, cross-over, placebo controlled and singleblinded in design. the effects of an oral intake of 150 mg pure EGCG over an acute period (2.5 hours), on key metabolic parameters and following an oral glucose challenge was measured (n = 8, healthy men). Results of this study were inconclusive on RMR, MST, CT plasma insulin, leptin and NEFA concentrations and appetite regulation. The second study tested the effects of varying doses (75, 150 and 300 mg/day) of pure EGCG on afore mentioned key metabolic parameters when supplemented over a period of 7 days each (n = 8, male). No significant changes in measured parameters were found. The results of these two studies indicate the need for further studies to investigate effects of EGCG on weight loss / weight maintenance possibly in combination with caffeine and/or other green tea flavanols. Both L-Histidine and EGCG supplmentations have previously shown to influence regulators of energy balance, blood glucose and cardiovascular risk factors via activation of the sympathetic nervous system in animal models. The present studies however did not provide any definitive conclusions following EGCG ingestion in healthy humans. L-Histidine supplementation brought about significant decrease in FBG and body weight after 10 days’ ingestion but its effect after 3.25 hours and 8 weeks was inconclusive. The effects following supplementation of both L-Histidine and EGCG ingestion needs to be further studied to clarify any potential metabolic consequences in humans.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Natural Drugs from Plants

Natural Drugs from Plants emphasizes the importance of medicinal plants for drug discovery worldwide. Chapters discuss the active ingredients of certain medicinal plants, their mechanisms of action, and how they can be used to treat different diseases

Inducción de compuestos con actividad antiinflamatoria en cultivos de células en suspensión de Sphaeralcea angustifolia (Cav.) G. Don (Malvaceae) a nivel de matraces y en biorreactor

Inflammation is a physiopathological process associated with a great number of chronic degenerative afflictions, mainly rheumatic, cardiovascular, gastric, and neurological. Rheumatic diseases are one of the principal health problems worldwide; in Mexico, persons at productive employment age are those most affected by these diseases. Thus, it is a priority health theme and one of economic importance that motivates the search for novel anti-inflammatory agents that act on therapeutic targets-of-interest. Medicinal plants are a great source of compounds that possess chemical structural diversity with biological potential; therefore, the former can be utilized as medicaments or as a base for the production of these. An important number of plant have been utilized traditionally for the treatment of patients with diseases that lead to an inflammatory process; among these, Sphaeralcea angustifolia is highlighted. This species has the pharmacological support that sustains its traditional use, as well as clinical evaluation in patients with osteoarthritis of a standardized phytomedicament in the content of the antiinflammatory, scopoletin. Although this, chemical studies on the species are scarce; thus, identifying and characterizing other compounds that contribute to the antiinflammatory effect of S. angustifolia are important. Considering that the plant grows in isolated populations and its collection is controlled by the Mexican Ministry of the Environment and Natural Resources (SEMARNAT, NOM-007-RECNAT-1997), it was indispensable to find alternatives for obtaining the plant tissue and conserving the species. For this purpose, cultures were developed of morphogenetic calluses by means of which we achieved indirect micropropagation of the plant. Similarly, the cultures of cells in suspension producers of the active scopoletin compound were established. This system was employed to evaluate a strategy of stimulation, modifying the carbon:nitrogen equilibrium on reducing the content of nitrate in the Murashige and Skoog medium (MS) to 2.74 mM. This condition did not affect the growth of the cellular suspension, favored the production of scopoletin, and stimulated the production of another two phenolic-type compounds. In this project, we initially proposed to isolate and identify the structure of the two compounds produced in the cellular suspension of S. angustifolia cultured in MS medium with nitrate restriction, as well as to evaluate their effect on the formation of plantar edema induced with λ-carrageenan and auricular edema with 12-oTetradecanoylphorbol-13-acetate (TPA). Nuclear magnetic resonance (NMR) spectroscopic data permitted the structural identification of tomentin (5-hydroxy-6,7- dimethoxycoumarin) and 2-(1,8-dihydroxy-4-isopropyl-6-methyl-7-methoxy) naphthoic acid, which was isolated and identified, to our knowledge for the first time, in cell cultures in suspension in a species of the Sphaeralcea genus, denominating this sphaeralcic acid. Both compounds inhibited the development of subplantar edema, an activity that was similar to that of the drug Indomethacin. In the auricular-edema model, the antiinflammatory activity of tomentin was similar to that of the synthetic drugs (Indomethacin and Naproxen), while that of the sphaeralcic acid (ED50 of 0.54 mg/ear) was significantly superior to the effect exhibited by the tomentin and the drug Naproxen. Later, this study was focused on optimizing the production of the active compounds (scopoletin, tomentin, and sphaeralcic acid) produced in the cellular suspensions of S. angustifolia, without taking into account the behavior of cellular growth, the latter one of the factors associated with production. Thus, we proposed establishing optimal levels of nitrate and copper, which stimulates the attack of a pathogenic organism, employing Surface Response Methodology (SRM) as model. During the Factorial Design (FD) stage, we determined that nitrate and copper levels in MS medium did not modify the speed of growth (0.317–0.389 days–1 ) or the duplication time (1.78–2.18 days) of the cellular suspension of S. angustifolia; notwithstanding this, the greatest cellular biomass (14.62 g/L) was obtained on increasing the nitrate content (2.74 mM) and decreasing that of copper (0.1 μM). The greatest production of coumarins and sphaeralcic acid was observed on days 2 and 4 of culture, favoring by means of the interaction of nitrate and copper; the values of 2.74 mM of nitrates and of 2 μM of copper were utilized as a base for establishing the Ascending design step (ADS). In the ADS model, the growth of the cellular suspension was reduced 74% in the cultures developed with the greatest contents of both nutrients. Likewise, we determined that the copper concentration higher to 5.68 μM in the MS medium inhibited growth in the cellular suspension, causing lysis and cell death. The production of coumarins and sphaeralcic acid generated an inflexion point of the curve and, in order to find the optimal conditions, we evaluated the effect of the nitrate and copper in short concentration intervals. For completing the quadratic model (Central Composite Design [CCD]) and to find the optimal conditions of nitrate and copper, it was necessary to add to the FD four additional points (the Star Design [SD]), taking as point of origin the nutritional conditions of 2.74 mM of nitrate and of 2 µM of copper. In the CCD model, we determined that modification of nitrate and copper in the MS culture medium favorably modulated the production of tomentin, scopoletin, and sphaeralcic acid in cellular cultures in the S. angustifolia suspension. The interaction of 2.42 mM of nitrate and 1.81 μM copper stimulates the greatest production of coumarins, registering levels of 236.45 µg/L in biomasses and of 4,136. 96 μg/L in the MS medium on day 2 of culture. The interaction was not determined of the factors in the production of the sphaeralcic acid, predicting that the best conditions for their excretion was obtained with 0.32 mM of copper-free nitrates on day 4 (5,258.12 μg/L) of culture, while its accumulation (756.91 μg/L) was favored by the presence of 2.26 μM of copper and 0.32 mM of nitrate. The production of coumarins in response to the stimulus is similar to that of a phytoalexin; on the other hand, the production of the sphaeralcic acid is characteristic of a constitutive secondary metabolite. Next, we evaluated whether the anti-inflammatory compounds obtained in the suspension of S. angustifolia at the flask level in the cellular culture developed in the laboratory bioreactor with mechanical shaking. The suspensions produced scopoletin and tomentin with a tendency similar to that registered at the flask level; on the other hand, the maximal productions of sphaeralcic acid were observed on days 2 and 4 of culture. In these cases, with respect to the production registered flask level, the accumulation was 20.2 and 78.1 times greater, and its excretion was 6.5 and 16.9 times superior, respectively.La inflamación es un proceso fisiopatológico asociado a un gran número de padecimientos crónico degenerativos, principalmente reumáticos, cardiovasculares, gástricos y neurológicos. Las enfermedades reumáticas son uno de los principales problemas de salud en el mundo; en México, las personas en edad productiva laboral son las más afectadas por estas enfermedades. Por lo anterior, es un tema prioritario de salud y de importancia económica que motiva a la búsqueda de nuevos agentes antiinflamatorios que actúen sobre los blancos terapéuticos de interés. Las plantas medicinales son una gran fuente de compuestos que poseen una diversidad estructural química con potencial biológico, por lo que pueden ser utilizadas como medicamentos o como base para su producción. Un número importante de especies vegetales han sido utilizadas tradicionalmente para el tratamiento de pacientes con enfermedades que conllevan un proceso inflamatorio, dentro de estas se destaca Sphaeralcea angustifolia. Esta especie cuenta con el respaldo farmacológico que sustenta el uso tradicional, así como la evaluación clínica en pacientes con osteoartritis de un fitomedicamento estandarizado en el contenido del antiinflamatorio escopoletina. No obstante, los estudios químicos de la especie son escasos, por lo que es importante el identificar y caracterizar a otros compuestos que coadyuven al efecto antiinflamatorio de S. angustifolia. Considerando que la planta crece en poblaciones aisladas y su colecta está controlada por la SEMARNAT (NOM-007-RECNAT-1997) fue indispensable búscar alternativas para la obtención de tejido vegetal y conservación de la especie. Para tal propósito, se desarrollaron cultivos de callos morfogenéticos a través de los cuales se logró la micropropagación indirecta de la planta. Asimismo, se establecieron cultivos de células en suspensión productores del compuesto activo escopoletina. Este sistema se empleó para evaluar una estrategia de estimulación modificando el balance carbono:nitrógeno, al reducir el contenido de nitrato en el medio de Murashige y Skoog (MS) a 2.74 mM; condición que no afectó el crecimiento de la suspensión celular, favoreció la producción de la escopoletina y estimuló la producción de otros dos compuestos del tipo fenólico. En este proyecto inicialmente se planteó aislar e identificar la estructura de los dos compuestos producidos en la suspensión celular de S. angustifolia cultivada en medio MS con restricción de nitrato, así como evaluar su efecto sobre la formación de los edemas plantar inducido con λ-carragenina y auricular con TPA (12-otetradecanoilforbol-13-acetato). Los datos espectroscópicos de Resonancia Magnética Nuclear (RMN) permitieron identificar estructuralmente a la tomentina (5-hidroxi-6,7- dimetoxicumarina) y al ácido 2-(1,8-dihidroxi-4-isopropil-6-metil-7-metoxi) naftoico, el cual se aisló e identificó por primera vez en cultivos de células en suspensión de una especie del género Sphaeralcea, denominándolo como ácido sphaerálcico. Ambos compuestos inhibieron el desarrollo del edema subplantar, actividad que fue similar a la del fármaco indometacina. En el modelo de edema auricular la actividad antiinflamatoria de la tomentina fue similar a la de los fármacos sintéticos (indometacina y naproxeno); en tanto que la del ácido sphaerálcico (DE50 de 0.54 mg/oreja) fue significativamente superior a el efecto mostrado por la tomentina y el fármaco naproxeno. Posteriormente, este estudio se enfocó en optimizar la producción de los compuestos activos (escopoletina, tomentina y ácido sphaerálcido) producidos en las suspensiones celulares de S. angustifolia, sin dejar de tomar en cuenta el comportamiento del crecimiento celular, siendo este uno de los factores asociado a la producción. Así, se planteó establecer los niveles óptimos de nitrato y de cobre, el cual simula el ataque de un microorganismo patógeno, empleando la metodología de superficie de respuesta (MSR) como modelo. En la etapa de Diseño Factorial (DF) se determinó que los niveles de nitrato y cobre en el medio MS no modificaron la velocidad de crecimiento (0.317-0.389 días-1 ) ni el tiempo de duplicación (1.78-2.18 días) de la suspensión celular de S. angustifolia; no obstante, la mayor biomasa celular (14.62 g/L) se obtuvo al incrementarse el contenido de nitrato (2.74 mM) y disminuyendo el del cobre (0.1 μM). La mayor producción de cumarinas y ácido sphaerálcico se observó a los 2 y 4 días de cultivo, favorecidos por la interacción de los contenidos de nitrato y cobre, los valores de 2.74 mM de nitratos y 2 μM de cobre fueron utilizados como base para el establecimiento del Diseño de Paso Ascendente (DPA). En el modelo DPA el crecimiento de los cultivos celulares se redujo en un 74% en los cultivos desarrollados con mayores contenidos de ambos nutrientes. Asimismo, se determinó que contenidos por arriba de 5.68 μM de cobre en el medio MS inhiben el crecimiento en la suspensión celular, provocando lisis y muerte celular. La producción de cumarinas y el ácido sphaerálcico, generó un punto de inflexión de la curva y para encontrar las condiciones óptimas se evaluó el efecto del nitrato y cobre en intervalos de concentración cortos. Para completar el modelo cuadrático (Diseño Compuesto Central, DCC) y encontrar las condiciones de nitrato y de cobre óptimas, fue necesario añadir al DF cuatro puntos más (Diseño en Estrella, DE) tomando como punto de origen las condiciones nutricionales de 2.74 mM de nitratos y 2 µM de cobre. En el modelo DCC se determinó que la modificación en la concentración de nitrato y cobre en el medio de cultivo MS moduló favorablemente la producción de tomentina, escopoletina y ácido sphaerálcico en los cultivos de células en suspensión de S. angustifolia. La interacción de 2.42 mM de nitratos y 1.81 μM de cobre estimula la mayor producción de las cumarinas, registrando niveles de 236.45 µg/L en biomasas y de 4,136. 96 μg/L en el medio a los 2 días de cultivo. No se determinó interacción de los factores sobre producción de ácido sphaerálcico, prediciendo que las mejores condiciones para su excreción se obtuvieron con 0.32 mM de nitratos libre de cobre a los 4 días (5,258.12 μg/L) de cultivo; mientras que su acumulación (756.91 μg/L) se favoreció con la presencia de 2.26 μM de cobre y 0.32 mM de nitrato. La producción de las cumarinas en respuesta al estímulo es similar a la de una fitoalexína, en cambio, la producción del ácido sphaerálcico es característica de un metabolito secundario constitutivo. A continuación, se evaluó si los compuestos antiinflamatorios obtenidos en la suspensión de S. angustifolia a nivel de matraces, se producen en el cultivo celular desarrollado en biorreactor de laboratorio con agitación mecánica. Las suspensiones produjeron escopoletina y tomentina con una tendencia similar a lo registrado a nivel de matraz; en cambio, las máximas producciones de ácido sphaerálcico se observaron a los 2 y 4 días de cultivo. En estos casos, con respecto a la producción registrada a nivel de matraz, la acumulación fue 20.2 y 78.1 veces mayor, y su excreción fue 6.5 y 16.9 veces superior, respectivamente