Angel Gil-Izquierdo

Betalains in the era of global agri-food science, technology and nutritional health

Natural pigments from plants are of growing interest as substitutes for synthetic dyes in the food and pharmaceutical industry and they increase their added value if they possess positive effects on health. These pigments can be added as such if they are in the legal authorized lists of additives or can be added as phytochemical-enriched plant extract achieving the original product, which has received it, the new nomenclature of functional food. In this way, we comprise on this review a wide point of view of a group of natural pigments known as betalains. From a chemical point of view, betalains are ammonium conjugates of betalamic acid with cyclo-DOPA (betacyanins, violet) and aminoacids or amines (betaxanthins, orange or yellow), which are compounds present in our diet. Besides and taking into account that one type of betalain, betanin is approved as food colorant (E-162) by the European Union and that enlarges the specific weight of these compounds in the diet, we have evolved an overview from the biosynthesis, technology and promoting production, industrial uses as pigments up to physiological and nutritional biovailability or biological and health-promoting properties of betalains for accessible information to industrials, researchers and consumers.

Occurrence of urolithins, gut microbiota ellagic acid metabolites and proliferation markers expression response in the human prostate gland upon consumption of walnuts and pomegranate juice

Epidemiology supports the important role of nutrition in prostate cancer (PCa) prevention. Pomegranate juice (PJ) exerts protective effects against PCa, mainly attributed to PJ ellagitannins (ETs). Our aim was to assess whether ETs or their metabolites ellagic acid and urolithins reach the human prostate upon consumption of ET-rich foods and to evaluate the effect on the expression of three proliferation biomarkers. Sixty-three patients with BPH or PCa were divided into controls and consumers of walnuts (35 g walnuts/day) or pomegranate (200 mL PJ/day) for 3 days before surgery. Independently of the ETs source, the main metabolite detected was urolithin A glucuronide, (3,8-dihydroxy-6H-dibenzo[b,d]pyran-6-one glucuronide) (up to 2 ng/g) together with the traces of urolithin B glucuronide, (3-hydroxy-6H-dibenzo[b,d]pyran-6-one glucuronide) and dimethyl ellagic acid. The small number of prostates containing metabolites was likely caused by clearance of the compounds during the fasting. This was corroborated in a parallel rat study and thus the presence of higher quantities of metabolites at earlier time points cannot be discarded. No apparent changes in the expression of CDKN1A, MKi-67 or c-Myc were found after consumption of the walnuts or PJ. Our results suggest that urolithin glucuronides and dimethyl ellagic acid may be the molecules responsible for the beneficial effects of PJ against PCa.

Identification of phenolic compounds in isolated vacuoles of the medicinal plant Catharanthus roseus and their interaction with vacuolar class III peroxidase: an H2O2 affair?

Class III peroxidases (Prxs) are plant enzymes capable of using H(2)O(2) to oxidize a range of plant secondary metabolites, notably phenolic compounds. These enzymes are localized in the cell wall or in the vacuole, which is a target for secondary metabolite accumulation, but very little is known about the function of vacuolar Prxs. Here, the physiological role of the main leaf vacuolar Prx of the medicinal plant Catharanthus roseus, CrPrx1, was further investigated namely by studying its capacity to oxidize co-localized phenolic substrates at the expense of H(2)O(2). LC-PAD-MS analysis of the phenols from isolated leaf vacuoles detected the presence of three caffeoylquinic acids and four flavonoids in this organelle. These phenols or similar compounds were shown to be good CrPrx1 substrates, and the CrPrx1-mediated oxidation of 5-O-caffeoylquinic acid was shown to form a co-operative regenerating cycle with ascorbic acid. Interestingly, more than 90% of total leaf Prx activity was localized in the vacuoles, associated to discrete spots of the tonoplast. Prx activity inside the vacuoles was estimated to be 1809 nkat ml(-1), which, together with the determined concentrations for the putative vacuolar phenolic substrates, indicate a very high H(2)O(2) scavenging capacity, up to 9 mM s(-1). Accordingly, high light conditions, known to increase H(2)O(2) production, induced both phenols and Prx levels. Therefore, it is proposed that the vacuolar couple Prx/secondary metabolites represent an important sink/buffer of H(2)O(2) in green plant cells.

Melatonin is synthesised by yeast during alcoholic fermentation in wines

Melatonin (N-acetyl-5-methoxytryptamine) is a neurohormone produced in the pineal gland. Its biological properties are related to the circadian rhythm. Recently, the European Food Safety Authority (EFSA) accepted the health claim related to melatonin and the alleviation of subjective feelings of jet lag. This molecule has been detected in some foods. In this work, 13 grape varieties were studied; 7 monovarietal wines were produced in an experimental winery under strictly controlled conditions and were sampled in different steps. The grape varieties used to make the wines were: Cabernet Sauvignon, Merlot, Syrah, Tempranillo, Tintilla de Rota, Palomino Fino and Alpha red. Liquid chromatography tandem mass spectrometry (LC-MS/MS) unequivocally confirmed the presence of melatonin in wines. The main contribution of this paper is the results that clearly show that melatonin is synthesised during the winemaking process, specifically after the alcoholic fermentation. Indeed, melatonin is absent in grapes and musts and is formed during alcoholic fermentation.

Hesperidin inhibits ovariectomized-induced osteopenia and shows differential effects on bone mass and strength in young and adult intact rats

The main aim of this study was to investigate the bone-sparing effect of hesperidin, one of the main flavonoid present in oranges, in two age groups of ovariectomized female rats, compared with their intact controls. Young (3 mo) and adult (6 mo) female Wistar rats were sham operated (SH) or ovariectomized (OVX) and then pair-fed for 90 days a casein-based diet supplemented or not with 0.5% hesperidin (Hp; n = 10/group). In older rats, Hp intake led to a partial inhibition of OVX-induced bone loss, whereas a complete inhibition was obtained in younger animals. At both ages, while plasma osteocalcin concentrations were unchanged, urinary excretion of deoxypyridinoline was reduced by Hp intake, suggesting that Hp was able to slow down bone resorption. Unexpectedly, in intact young rats, Hp consumption resulted in a significant increase in bone mineral density (BMD). Indeed, 6-mo-old HpSH rats had a similar BMD to 9-mo-old nontreated SH adult rats, suggesting an accelerated bone mass gain in the young rats. In contrast, in intact adult rats, Hp did not further increase BMD but did improve their bone strength. The results of this study show a protective effect of Hp on bone loss in OVX rats of both ages without uterine stimulation and accompanied by a lipid-lowering effect. The unexpected and intriguing findings obtained in intact rats showing improved BMD in young rats and improved femoral load in adult rats merit further investigation. The bone and lipid benefits of hesperidin make it an attractive dietary agent for the management of the health of postmenopausal women.

In vitro studies to assess the antidiabetic, anti-cholinesterase and antioxidant potential of Spergularia rubra

Spergularia rubra is distributed all over the world, being its infusion used as diuretic. In spite of its large use, the antidiabetic, anti-cholinesterase and antioxidant activities of this species have not been assessed and its chemical composition is scarcely known. In the work herein a hydromethanolic extract was studied. Thirty-six phenolic compounds were determined by HPLC-DAD, comprising non-acylated C-glycosyl flavones (38%), C-glycosyl flavones acylated with aromatic acids (36%), C-glycosyl flavones acylated with aliphatic acids (13%) and 10% corresponded to C-glycosyl flavones with a mixed acylation. Organic acids (oxalic, citric, malic, quinic and fumaric acids) and fatty acids (azelaic, myristic, palmitic, linoleic, linolenic and stearic acids) are described for the first time. Their determination by HPLC-UV and GC-IT-MS allowed finding concentrations of 192.15 and 34.87g/kg, respectively. The extract showed a dose-dependent response against DPPH, superoxide and nitric oxide radicals. The same effect was observed in the α-glucosidase inhibitory assay and against acetylcholinesterase and butyrylcholinesterases. The bioactivities observed may be due, at least partially, to the presence of the different metabolites determined in the present study. The results suggest that the dried extract of S. rubra may be interesting for incorporation in pharmaceutical preparations for human health, since it can suppress hyperglycaemia and inhibit cholinesterases, and or as food additive due to its antiradical activity.

Bauhinia forficata Link authenticity using flavonoids profile: Relation with their biological properties

HPLC-DAD-ESI/MS(n) was used to ascertain the authenticity of two certified and two commercial Bauhinia forficata Link samples. Different flavonoids profiles were obtained, involving 39 compounds. Just kaempferol-3-O-(2-rhamnosyl)rutinoside was found in all analysed samples. Five compounds were common to the certified samples of B. forficata Link and B. forficata Link subsp. pruinosa (Vogel) Fortunato & Wunderlin, being kaempferol derivatives the most representative ones. The phenolic composition of B. forficata Link subsp. pruinosa (Vogel) Fortunato & Wunderlin is described herein for the first time, accounting for eight compounds, while 10 new compounds were identified in B. forficata Link. Commercial B. forficata Link showed higher contents of quercetin derivatives, in addition to the presence of myricetin derivatives and flavonoids-(galloyl)glycosides, for which the MS fragmentation pattern is reported for the first time. B. forficata Link and the two commercial samples were able to inhibit α-glucosidase, with EC(50) values lower than that found for acarbose. Mild effects on cholinesterases were observed with the certified samples, while commercial ones were more effective. The same behaviour was observed concerning the scavenging of DPPH, nitric oxide and superoxide radicals. The presence of high contents of quercetin derivatives in commercial samples seems to directly influence their biological properties. The differences between phenolic profiles and their relation with the authenticity of commercial samples are discussed.

Increased bioavailability of hesperetin-7-glucoside compared with hesperidin results in more efficient prevention of bone loss in adult ovariectomised rats.

Hesperidin (Hp), a citrus flavonoid predominantly found in oranges, shows bone-sparing effects in ovariectomised (OVX) animals. In human subjects, the bioavailability of Hp can be improved by the removal of the rhamnose group to yield hesperetin-7-glucoside (H-7-glc). The aim of the present work was to test whether H-7-glc was more bioavailable and therefore more effective than Hp in the prevention of bone loss in the OVX rat. Adult 6-month-old female Wistar rats were sham operated or OVX, then pair fed for 90 d a casein-based diet supplemented or not with freeze-dried orange juice enriched with Hp or H-7-glc at two dose equivalents of the hesperetin aglycone (0.25 and 0.5 %). In the rats fed 0.5 %, a reduction in OVX-induced bone loss was observed regarding total bone mineral density (BMD):+7.0 % in OVX rats treated with Hp (HpOVX) and +6.6 % in OVX rats treated with H-7-glc (H-7-glcOVX) v. OVX controls (P < 0.05). In the rats fed 0.25 % hesperetin equivalents, the H-7-glcOVX group showed a 6.6 % improvement in total femoral BMD v. the OVX controls (P < 0.05), whereas the Hp diet had no effect at this dose. The BMD of rats fed 0.25 % H-7-glc was equal to that of those given 0.5 % Hp, but was not further increased at 0.5 % H-7-glc. Plasma hesperetin levels and relative urinary excretion were significantly enhanced in the H-7-glc v. Hp groups, and the metabolite profile showed the absence of eriodictyol metabolites and increased levels of hesperetin sulphates. Taken together, improved bioavailability of H-7-glc may explain the more efficient bone protection of this compound.

The effect of storage temperatures on vitamin C and phenolics content of artichoke (Cynara scolymus L.) heads

Abstract The effect of temperatures (0, 2, 5, 7 and 10°C) for 14 days of storage on antioxidant constituents such as vitamin C and phenolic compounds of artichoke c.v. ‘Blanca de Tudela’ was studied. Three groups of phenolics were identified and quantified as: chlorogenic acid; 1,5-dicaffeoylquinic acid (1,5 di-CQA)+3,5-dicaffeoylquinic acid (3,5 di-CQA); and 1,4-dicaffeoylquinic acid (1,4-di-CQA)+4,5-dicaffeoylquinic acid (4,5 di-CQA). The content of vitamin C was higher in the internal bracts compared with the external ones (144 and 193 mg kg −1 fresh wt., respectively) and decreased after 14 days of storage under the assayed temperatures. Each individual group of phenolics was 10-fold higher in the internal bracts than in the external ones. At harvest, total phenolics (618 mg kg −1 ), chlorogenic acid (143 mg kg −1 ) and 1,4 di-CQA+4,5 di-CQA (207 mg kg −1 ), increased in internal bracts after storage, particularly at 2, 5 and 7°C. On the other hand, 1,5-di-CQA+3,5-di-CQA decreased from 260 to 150 mg kg −1 after storage. Consumption of the edible artichoke fraction could be an important source of natural antioxidants and provide significant nutritional value to the diet.

Differential effects of two citrus flavanones on bone quality in senescent male rats in relation to their bioavailability and metabolism.

The effect of hesperidin (Hp) and naringin (Nar), two major citrus flavanones, on the regulation of bone metabolism was examined in male senescent rats. Twenty -month -old gonad-intact male Wistar rats received a casein-based diet supplemented with or without either 0.5% hesperidin (Hp), 0.5% naringin (Nar) or a mix of both flavanones (Hp+Nar, 0.25% each). After 3 months, daily Hp intake significantly improved femoral bone integrity as reflected by improvements in total and regional bone mineral densities (BMD) (9.7%-12.3% improvements, p<0.05) and trabecular bone volume fraction (24.3% improvement, p<0.05) at the femur compared with control group. In contrast, naringin exerted site-specific effects on BMD (10.2% improvement at the distal metaphyseal area, p<0.05) and no further benefit to bone mass was observed with the mix of flavanones. Bone resorption (DPD) was significantly attenuated by Hp and Nar given alone (40.3% and 26.8% lower compared to control, p<0.05, respectively) but not by the mixture of the two. All treatments significantly reduced expression of inflammatory markers to a similar extent (IL-6, 81.0-87.9% reduction; NO, 34.7-39.5% reduction) compared to control. Bone formation did not appear to be strongly affected by any of the treatments (no effect on osteocalcin levels, modest modulation of tibial BMP-2 mRNA). However, as previously reported, plasma lipid-lowering effects were observed with Hp and Nar alone (34.1%-45.1% lower for total cholesterol and triglycerides compared to control, p<0.05) or together (46% lower for triglycerides, p<0.05). Surprisingly the plasma circulating level of naringin (8.15μM) was >5-fold higher than that of hesperidin (1.44μM) at equivalent doses (0.5%) and a linear reduction in plasma levels was observed upon co-administration (0.25% each) indicating absence of competition for their intestinal absorption sites and metabolism. The higher efficacy of Hp at a lower plasma concentration than naringin, as well as the identification of the major circulating metabolite of hesperidin (hesperetin-7-O-glucuronide) underlines the importance of flavanone bioavailability and metabolism in their biological efficacy and suggests a structure-function relationship in the mechanism of action of the active metabolites.