This document describes a study that developed a high performance liquid chromatography coupled to mass spectrometry (HPLC-MS/MS) method for profiling and quantifying folate monoglutamates in tomato fruits. Several parameters of the folate extraction process from tomato tissues were optimized, including extraction conditions, pH range, amount of tri-enzyme used, and boiling time. The extracted samples were then purified using ultra-filtration and analyzed using HPLC-MS/MS. The method was validated for linearity, sensitivity and recovery. This HPLC-MS/MS method was then applied to quantify folates in other plants like spinach, capsicum and garden pea, demonstrating its versatility for accurate determination of different folate
Influence of Ethanolic Extractives of Leaves of Mulberry, Morus Alba (L) On 7...iosrjce
The study deals with investigation of thechemopreventive potential and antilipidperoxidative effects
of ethanolic leaf extract of mulberry, Morus alba (L) (TpEt) on 7,12-dimethylbenz(a)anthracene (DMBA)-
induced buccal pouch carcinoma in Syrian hamster, Mesocricetusauratus (L). Oral squamous cell carcinoma
was developed in the buccal pouch of Syrian golden hamsters, by painting with 0.5% DMBA in liquid paraffin,
thrice a week, for 14 weeks. The tumor incidence, volume and burden were determined. Oral administration
ofTpEt at a dose of 300 mg/kg, body weight, to DMBA (on alternate days for 14 weeks)- painted animals
significantly prevented the incidence, volume and burden of the tumor. TpEt showed potent antilipidperoxidative
effect, as well as enhanced the antioxidant status in DMBA- painted animals. TpEt has potent chemopreventive
efficacy and significant antilipidperoxidative effect, in DMBA-induced oral carcinogenesis. The active principle
of mulberry leaf may have the abilities of induction of apoptosis, which involve disruption of mitochondrial
membrane potential, release of cytochrome C and activation of caspase. With it’s bioactive compounds,
mulberry, Morus alba (L) may open a new avenue in the cancer prevention and treatment.
The results of this study revealed that the use of natural bees’ honey has the ability to protect the liver of rats against the toxic effects of melamine.
Melamine contains 66% nitrogen by mass, so it is sometimes illegally added to food products in order to increase the apparent protein content that has recently become a serious concern. The illegal use of melamine as a food ingredient has led to many poisoning incidents of cats and dogs in the United States, as well as renal function failure of Chinese infants.
Extraction and charcaterization of beta glucan from oat for industrial utiliz...Dr Asif Ahmad
This document summarizes an article that appeared in the International Journal of Biological Macromolecules. The article studied the extraction and characterization of beta-D-glucan from oat for industrial utilization. It analyzed the effect of different extraction methods on the yield, chemical composition, and functional properties of extracted beta-D-glucan gum pellets. The highest yield of 5.14% was obtained using an enzymatic extraction process. Chemical analysis showed protein as the main impurity. The extracted beta-D-glucan exhibited good water binding capacity, viscosity, color properties, and foaming stability suitable for industrial applications.
Extraction of beta glucan from oat and its lipoprotein profileDr Asif Ahmad
This document summarizes a study that extracted β-glucan from oat at various temperatures and pH levels. Higher extraction temperatures and neutral pH levels increased the yield of gum pellet extracted and the recovery of β-glucan. Extraction at 50°C and pH 7 was found to extract the highest amounts of soluble and total dietary fiber, while removing more impurities from the gum pellet. Rats fed diets containing the extracted gum pellet at this condition showed reductions in serum glucose, total cholesterol, triglycerides, and LDL cholesterol levels compared to controls.
Quantifying phytate in dairy digesta and feces- Alkaline extraction and high-...Partha Ray
This document describes the development of a method to accurately quantify undigested phytate in ruminant feces and digesta. Samples were collected from dairy heifers and cows fed diets varying in phytate content to challenge extraction and quantification methods. Alkaline extraction using NaOH and EDTA yielded greater estimates of phytate in digesta and fecal samples than acid extraction, but alkaline extracts cannot be directly analyzed by high-performance ion chromatography (HPIC) due to matrix interference. The proposed method combines the extraction power of alkaline extraction with the precision of HPIC analysis by acidifying alkaline extracts and filtering to remove matrix interference before HPIC quantification. Validation tests found near complete recovery of added phyt
Nutritional Profile and Physicochemical Properties of Peach Varieties in Ethi...AJSERJournal
Nutritional and functional characteristics of fruits are related to their quality and are influenced by
genotype and ripening stage, and by environmental conditions and orchard management practices. The purpose of this
research was to test nutritional profile and selected physicochemical properties of different improved and adopted
peach varieties in Ethiopia and comparative study among varieties and between study varieties and standard
reference. The result got show that improved peach varieties greater amount of ash (4.3-5.51%), protein (4.34-6.05%),
fat (0.097-1.386%) and fiber (2.87-4.611%) than standard reference (0.263, 1.423, 0.427, and 1.20%, in the
aforementioned order) but lower in carbohydrate (84.411-89.90%) than standard of (96.678%). The mineral content
higher in K (0.48-1.182%), Ca (0.162-0.565 %), Mg (0.037-0.066%), Fe (16.33-159.2%) and Zn (2.43-8.84%) than
standard reference (0.942, 0.051, 0.071, 16.32, 3.213%), respectively except Na, Mg, Cu and Sulfur while the fruit
quality was revealed low moisture content which less water and medium vitamin C (3.99-5.55%) and TSS (8.36-
14.31%). We observed variation in proximate composition and mineral content among sixteen peach varieties.
Exploring the potential of red kidney beans Dr Asif Ahmad
This study explored the potential of red kidney beans to develop a protein-based food product. The beans were found to contain 25.78% protein as well as carbohydrates, fiber, fat, ash, and various minerals. A protein isolate was prepared from the beans and its functional properties including solubility, emulsification, gelation, and foaming were evaluated at different pH levels. The solubility, emulsifying activity, foam capacity, and stability were dependent on pH, with minimum values observed around pH 4-5 and maximum values around pH 10. Gelation properties improved at acidic pH. Thus, red kidney bean protein isolate has functional properties that can be modified by pH for use in various food formulations.
Influence of Ethanolic Extractives of Leaves of Mulberry, Morus Alba (L) On 7...iosrjce
The study deals with investigation of thechemopreventive potential and antilipidperoxidative effects
of ethanolic leaf extract of mulberry, Morus alba (L) (TpEt) on 7,12-dimethylbenz(a)anthracene (DMBA)-
induced buccal pouch carcinoma in Syrian hamster, Mesocricetusauratus (L). Oral squamous cell carcinoma
was developed in the buccal pouch of Syrian golden hamsters, by painting with 0.5% DMBA in liquid paraffin,
thrice a week, for 14 weeks. The tumor incidence, volume and burden were determined. Oral administration
ofTpEt at a dose of 300 mg/kg, body weight, to DMBA (on alternate days for 14 weeks)- painted animals
significantly prevented the incidence, volume and burden of the tumor. TpEt showed potent antilipidperoxidative
effect, as well as enhanced the antioxidant status in DMBA- painted animals. TpEt has potent chemopreventive
efficacy and significant antilipidperoxidative effect, in DMBA-induced oral carcinogenesis. The active principle
of mulberry leaf may have the abilities of induction of apoptosis, which involve disruption of mitochondrial
membrane potential, release of cytochrome C and activation of caspase. With it’s bioactive compounds,
mulberry, Morus alba (L) may open a new avenue in the cancer prevention and treatment.
The results of this study revealed that the use of natural bees’ honey has the ability to protect the liver of rats against the toxic effects of melamine.
Melamine contains 66% nitrogen by mass, so it is sometimes illegally added to food products in order to increase the apparent protein content that has recently become a serious concern. The illegal use of melamine as a food ingredient has led to many poisoning incidents of cats and dogs in the United States, as well as renal function failure of Chinese infants.
Extraction and charcaterization of beta glucan from oat for industrial utiliz...Dr Asif Ahmad
This document summarizes an article that appeared in the International Journal of Biological Macromolecules. The article studied the extraction and characterization of beta-D-glucan from oat for industrial utilization. It analyzed the effect of different extraction methods on the yield, chemical composition, and functional properties of extracted beta-D-glucan gum pellets. The highest yield of 5.14% was obtained using an enzymatic extraction process. Chemical analysis showed protein as the main impurity. The extracted beta-D-glucan exhibited good water binding capacity, viscosity, color properties, and foaming stability suitable for industrial applications.
Extraction of beta glucan from oat and its lipoprotein profileDr Asif Ahmad
This document summarizes a study that extracted β-glucan from oat at various temperatures and pH levels. Higher extraction temperatures and neutral pH levels increased the yield of gum pellet extracted and the recovery of β-glucan. Extraction at 50°C and pH 7 was found to extract the highest amounts of soluble and total dietary fiber, while removing more impurities from the gum pellet. Rats fed diets containing the extracted gum pellet at this condition showed reductions in serum glucose, total cholesterol, triglycerides, and LDL cholesterol levels compared to controls.
Quantifying phytate in dairy digesta and feces- Alkaline extraction and high-...Partha Ray
This document describes the development of a method to accurately quantify undigested phytate in ruminant feces and digesta. Samples were collected from dairy heifers and cows fed diets varying in phytate content to challenge extraction and quantification methods. Alkaline extraction using NaOH and EDTA yielded greater estimates of phytate in digesta and fecal samples than acid extraction, but alkaline extracts cannot be directly analyzed by high-performance ion chromatography (HPIC) due to matrix interference. The proposed method combines the extraction power of alkaline extraction with the precision of HPIC analysis by acidifying alkaline extracts and filtering to remove matrix interference before HPIC quantification. Validation tests found near complete recovery of added phyt
Nutritional Profile and Physicochemical Properties of Peach Varieties in Ethi...AJSERJournal
Nutritional and functional characteristics of fruits are related to their quality and are influenced by
genotype and ripening stage, and by environmental conditions and orchard management practices. The purpose of this
research was to test nutritional profile and selected physicochemical properties of different improved and adopted
peach varieties in Ethiopia and comparative study among varieties and between study varieties and standard
reference. The result got show that improved peach varieties greater amount of ash (4.3-5.51%), protein (4.34-6.05%),
fat (0.097-1.386%) and fiber (2.87-4.611%) than standard reference (0.263, 1.423, 0.427, and 1.20%, in the
aforementioned order) but lower in carbohydrate (84.411-89.90%) than standard of (96.678%). The mineral content
higher in K (0.48-1.182%), Ca (0.162-0.565 %), Mg (0.037-0.066%), Fe (16.33-159.2%) and Zn (2.43-8.84%) than
standard reference (0.942, 0.051, 0.071, 16.32, 3.213%), respectively except Na, Mg, Cu and Sulfur while the fruit
quality was revealed low moisture content which less water and medium vitamin C (3.99-5.55%) and TSS (8.36-
14.31%). We observed variation in proximate composition and mineral content among sixteen peach varieties.
Exploring the potential of red kidney beans Dr Asif Ahmad
This study explored the potential of red kidney beans to develop a protein-based food product. The beans were found to contain 25.78% protein as well as carbohydrates, fiber, fat, ash, and various minerals. A protein isolate was prepared from the beans and its functional properties including solubility, emulsification, gelation, and foaming were evaluated at different pH levels. The solubility, emulsifying activity, foam capacity, and stability were dependent on pH, with minimum values observed around pH 4-5 and maximum values around pH 10. Gelation properties improved at acidic pH. Thus, red kidney bean protein isolate has functional properties that can be modified by pH for use in various food formulations.
Effects of Metformin, Pioglitazone and Aqueous Extract of Delonix Regia on Bl...iosrjce
The effects of Delonix regia extract (d200mg, d300mg, and d400mg), metformin (m8.3mg, m12.5mg
and m16.5mg), pioglitazone (p0.5mg, p0.7mg and p0.9mg) and combined formulation of metformin and extract
(m6.25d150mg) on glycated hemoglobin status in streptozotocin-induced diabetic Albino wistar rats. Diabetic
status of these rats was assessed by estimating fasting blood glucose levels. A total of 150 albino rats were used
for the investigation and were grouped into twelve groups of twelve rats each as follows; Group I: normal
control rats (NCR). Group II: Diabetic control rats (DCR). Group III: Diabetic rats treated with d200mg.
Group IV: Diabetic rats treated with d300mg. Group V: Diabetic rats treated with d400mg. Group VI: Diabetic
rats treated with m8.3mg. Group VII: Diabetic rats treated with m12.5mg. Group VIII: Diabetic rats treated
with m16.5mg. Group IX: Diabetic rats treated with p0.5mg. Group X: Diabetic rats treated with p0.75mg.
Group XI: Diabetic rats treated with p1.0mg. Group XII: Diabetic rats treated with m125d300mg each for male
and female respectively, for a total of 56 days. After every two weeks interval of treatment for eight weeks three
rats from each group were sacrificed and blood sample were collected and analyzed for various parameters.
The result obtained showed an elevated level of glycated hemoglobin in diabetic-induced wistar albino rats
compared with normal control rats. However, there was reversal of the effects when treated with the
drug/extract. Also there was reduction in the blood glucose level of the diabetic rats treated with metformin
(from 6.37±0.69 to 5.20±0.62mmol/l), pioglitazone (from 7.30±0.21mmol/l to 4.70±0.46), aqueous extract of
Delonixregia (from 8.20±0.81mmol/l to 6.10±0.60) and combined formulation of metformin and extract (from
7.81±0.34 to 4.80±0.17), at p<0.05 confidence level when compared with diabetic control rats in the various
weeks of treatment respectively
This study was designed to evaluate the effect of
70% ethanolic crude extract of Portulaca oleracea L on mice
orgons . (In vivo),In vivo, the acute toxicity of 70 % ethanolic
extract of the plant on normal mice was studied. No toxic effect
was noted on normal mice even at 9500 mg /kg B.W S/C
injection.Histopathological changes due to ethanolic extract of
the plant in healthy mice were summarized in hyperplasia of
white pulp with amyloid deposition, proliferation of
megakaryocytes and mononuclear cell infiltration in the liver and
kidney parenchyma. There were no significant lesions detected in
the brain, heart and ovary in all treated groups.
This document discusses using metabolomics to analyze the food metabolome - all metabolites derived from digestion and metabolism of food components. It notes the complexity of nutritional exposures from foods, which contain nutrients, non-nutrients from natural and man-made sources, and contaminants. Individual metabolic capacity is also influenced by genetics, microbiota, age, and other factors. The food metabolome is defined and applications are described, including discovering new food bioactives, understanding diet-health relationships, and personalized nutrition. Challenges in analyzing the large and diverse food metabolome are also outlined. Initial studies using targeted and untargeted metabolomics are discovering new biomarkers of food intake from various foods. Larger cohort studies and controlled interventions are needed
This study evaluated changes in bioactive compounds and antioxidant activity in the pulp and seeds of Bunchosia glandulifera fruit during different ripening stages. Key findings:
1) Levels of phenolic compounds, carotenoids, vitamin C, and caffeine increased from the second ripening stage to the ripe fruit stage.
2) Ripe fruit had the highest levels of bioactive compounds compared to earlier stages.
3) Phenolic compound and carotenoid levels were highest in the pulp.
4) Carotenoid content was positively correlated with antioxidant activity.
Biochemical effect of Aqueous Carica papaya Seed and Leaf Extracts on Serum B...iosrjce
The cost of administering modern anti diabetic drugs is usually beyond the reach of most people in
the low income group especially those in the developing world where the disease is on the increase. This has led
to the current shift to the use of preparations from various parts of plants because of the current increase in the
knowledge of their toxicity, side effects, active constituents and doses. This study was conducted to compare the
effects of the aqueous Carica papaya seed and leaf extracts on serum biochemistry of alloxan induced diabetic
rats especially glucose level. Male Wister rats weighing 150-200g were induced with single freshly prepared
alloxan monohydrate (150 mg/kg body weight).Diabetes was confirmed after seven days in alloxan-induced rats
showing fasting blood glucose levels ≥ 200mg/dl. The diabetic rats were randomly allocated into three
experimental groups which received Seed extract, leaf extract or normal saline depending on the group. The
extracts were administered orally for twenty-eight days after which the animals were sacrificed and blood
samples were collected for Biochemical analyses. The results showed that both extracts have significant
hypoglycaemic, hepatoprotective and nephroprotective effects although extract of the seed proved to be more
potent than that of the leaf
This document provides an overview of hepatoprotective activity and agents. It discusses the anatomy and functions of the liver, common liver disorders, and mechanisms of hepatotoxicity. Screening methods for evaluating hepatoprotective effects include in vitro, ex vivo, and in vivo models. Several plants are described that have shown hepatoprotective properties, including Picrorrhiza kurroa, Orthosiphon stamineus, Tridax procumbens, Boerhaavia diffusa, and Hoslundia opposita. The document provides references for further information.
Extraction of Colorant From Dragon Fruit Peel and It's Application in a baker...Salemir Hossain
This is a project or thesis proposal presentation for B.Sc Engineering level on Extraction of Colorant From Dragon Fruit Peel and It's Application in a bakery Product. If any help is needed then contact on salemirhossainjust@gmail.com
Diabetes mellitus was induced in rats using streptozotocin. Creatinine, urea, and inflammatory biomarkers were measured in the rats' blood serum. Creatinine and urea levels were significantly increased in diabetic rats, while an inflammatory marker (IL-10) decreased. Administration of Jatropha curcas extracts to diabetic rats resulted in reductions in creatinine, urea, and inflammatory biomarkers, similar to effects of the antidiabetic drug glibenclamide. The extracts showed protective effects against kidney disorders associated with diabetes mellitus.
The document summarizes a study that analyzed samples collected from diarrhea patients and their mothers in rural Bangladesh to detect coliform bacteria. Several species of coliform bacteria were isolated from the samples, with isolation rates ranging from 38.01-3.51%. Extracts of Moringa oleifera leaves were tested against the isolated bacteria. The leaf extracts showed antibacterial effects and were able to inhibit the growth of all tested bacterial pathogens, including E. coli, Shigella, Salmonella, Enterobacter, Klebsiella, and Serratia species. The extracts had zone of inhibition ranging from 8-23 mm and minimum inhibitory concentration values between 62.5-1000 μg/mL, suggesting M. oleifera extracts could
Fruit and Vegetable Waste Hydrolysates as Growth Medium for Higher Biomass an...Premier Publishers
Fruit and vegetable wastes include peels, pulp and seeds that constitute about 40% of the total mass and constitute huge environmental problems. Cultivation of microalgae that utilizes fruit and vegetable wastes as feedstock to produce value added products such as biomass and lipids is a unique approach. Different concentrations of fruit waste hydrolysate (FWH) and vegetable waste hydrolysate (VWH) were used for heterotropic cultivation of Chlorella vulgaris thereby optimizing the suitable hydrolysate concentration for higher biomass and lipid production. FWH in the ratio of 8:2 has produced maximum specific growth rate of 1.92 µ d-1. Higher biomass was recorded in growth medium supplemented with FWH (0.16 mg L-1) than VWH medium. Highest chlorophyll content of 7.2 mg L-1 was observed in 8:2 ratio of FWH whereas it was 4.3 mg L-1 in VWH at the same concentration. Carotenoid content was highest in VWH than FWH media with a maximum content of 0.52 and 0.42 mg L-1 respectively. Fruit waste hydrolysates significantly increased the total lipid content than the vegetable waste hydrolysate medium. Highest lipid content of 6.63 mg L-1 was recorded in 8:2 ratio of FWH. This work demonstrates the feasibility of fruit waste hydrolysate as a nutrient source for algal cultivation and a cost reduction of growth medium in algal biomass and lipid production.
Proximate and Toxicological Analyses of Detoxified Jatropha Curcas Seedsiosrjce
The need for detoxification of less utilized crops with similar nutritional composition with soybeans
as potential substitute or supplement for soybeans in conventional feeds production is obvious as price of
soybean continue to rise. Jatropha curcas is one of such crops which can serve as a potential source of dietary
energy and protein. However, the presence of anti-nutritional factors restricts the utilization of the Jatropha
curcas seed in animal feed. Several researchers however have shown that this obstacle can be overcome by
detoxifying the seeds, but many of them failed to established the effects of these detoxification methods on the
nutritional content of Jatropha curcas .The main objective of this study therefore was to determine the effect of
three simple inexpensive physical methods of detoxification (soaking, roasting and fermentation) on the
proximate and toxicological compositions of Jatropha curcas seed meal. To achieve this, Jatropha curcas seeds
sample used were divided into four parts. The first three parts were subjected to the three different physical
treatments after which they were dried to constant weight and while the fourth part was dried to constant weight
and milled. These four samples were then analysed for their proximate and toxicological composition .The
results showed that fermentation deactivated the antinutrients most in the seeds and did not adversely affect the
nutritional composition of the seeds
This research article evaluated the potential aphrodisiac effects of Moringa oleifera seed extracts in male albino rats. Researchers administered aqueous, alcohol, and chloroform extracts of M. oleifera seeds to male rats for 21 days. They observed several parameters of sexual behavior and found that the extracts significantly increased mounting frequency, intromission frequency, and ejaculation latency, while decreasing mounting latency, intromission latency, and post-ejaculatory interval. The extracts also increased libido without any adverse effects. The results suggest that M. oleifera seed extracts can enhance sexual behavior in male rats, supporting its traditional use as an aphrodisiac.
The International Journal of Engineering and Science (The IJES)theijes
The International Journal of Engineering & Science is aimed at providing a platform for researchers, engineers, scientists, or educators to publish their original research results, to exchange new ideas, to disseminate information in innovative designs, engineering experiences and technological skills. It is also the Journal's objective to promote engineering and technology education. All papers submitted to the Journal will be blind peer-reviewed. Only original articles will be published.
Phytochemical, Proximate and Nutrient Analysis of Cassia Tora Seedsinventionjournals
Recently Proximate analysis and phytochemical analysis and nutrient analysis of seeds of Cassia tora L. from the campus of Government Vidharbha Institute of Science and Humanity, Amravati had been investigated. The seeds sample contained tannin, saponin, protein, steriods, terpenoids, carbohydrate, alkaloids, flavonoids and glycosides. Proximate analysis of moisture, ash, fat amd mineral analysis of calcium, magnesium, iron, nitrogen and solubility were check. The values of it is moisture (56%), cold water (52%), hot water (54%), 1%NaOH (43%), 1%HCl (61%), benzene +alcohol (35%), ash content (17%). These results indicate that the seeds of these Cassia tora L. contains mineral and nutrients elements that will be useful in nutrition. Also the existence of some phytochemicals like tannin, saponin and steroids illustrated medicinal action of the plant in its therapeutic uses. The result of their phytochemcial screening could justify the observed activities and validate their use in herbal medicine.
This project report summarizes research conducted at the Central Science Laboratory (CSL) in the UK on assessing genetic variability and purifying bioactive proteins from oilseed rape (OSR) meal. The research was jointly funded by the European Commission and Home Grown Cereals Authority (HGCA) as part of a larger EU project involving 14 partners.
The studies at CSL showed that DNA fingerprinting, particularly microsatellite analysis, could be used to estimate genetic variability in protein composition among OSR varieties. Research also involved isolating and characterizing antifungal and proteinase inhibitor proteins from OSR meal that have potential applications in plant protection.
The overall EU project aimed to explore value-added uses for OS
Protective Effect of Leaves of Ficus carica Against Carbon Tetrachloride-Indu...UKJPB Journal
The document summarizes a study that evaluated the hepatoprotective effects of leaves of Ficus carica (fig leaves) against carbon tetrachloride (CCl4)-induced liver damage in rats. Various extracts (petroleum ether, ethyl acetate, methanol) of F. carica leaves were prepared and screened for phytochemicals. The ethyl acetate extract was evaluated for its ability to reduce CCl4-induced increases in serum markers of liver damage (SGOT, SGPT, bilirubin) and protect against histological liver damage. Results showed the ethyl acetate extract significantly reduced the elevated serum markers and reversed histological liver changes caused by CCl4, demonstrating hepatoprotective effects
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
All manuscripts are subject to rapid peer review. Those of high quality (not previously published and not under consideration for publication in another journal) will be published without delay.
Thyroid hormone (T3 and T4) deficiency on the metabolism of vitamin Aiosrphr_editor
Both cabbage and thiouracil exert high impact on thyroid hormone. In the present study it is found that decreasing amount of thyroid hormone (T3&T4) inhibit the metabolism of vitamin A in the freshwater fish Ctepharyngodon idella. The goal of this study was to determine the effect of deficiency of thyroid hormone on the retinoid reserve of the fish due to the administration of cabbage and thiouracil as feed. For this experiment fish Ctepharyngodon idella was kept in different feeding states. First was kept with aquatic weed hydrilla, mass grass, filamentous algae. Second group was kept with chopped cabbage and third group was feed with chopped cabbage along with thiouracil. Blood samples were collected from the caudal vasculature in the heparin zed syringes and plasma was assayed for plasma T3&T4 concentration by Radioimmunoassay(RIA). The vitamin A concentration i.e., retinol, dehydroretinol and β-carotene are estimated from the liver of the fish through HPLC procedure. The result shows concentration of mean plasma thyroid hormone was significantly decreased in fishes subjected to cabbage and thiouracil in their feed compared to those of fishes in controlled condition. It is also seen that retinol, dehydroretinol and β-carotene concentration also decreased to a considerable amount in fishes subjected to cabbage and thiouracil in their feed. So, from the present experiment, it is clear that thyroid deficiency can affect the retinoids reserves of fish.
1. Methyl donors such as folate, betaine, choline, and methionine are important nutrients that provide methyl groups for critical cellular processes like DNA methylation and protein synthesis.
2. Betaine donates methyl groups to homocysteine via the betaine homocysteine methyltransferase (BHMT) pathway in the liver and kidneys, the major organs involved in methyl group metabolism. This pathway is regulated by factors like S-adenosylmethionine.
3. Methyl donor deficiencies can disturb energy metabolism and protein synthesis in the liver and cause fatty liver or muscle disorders. Elevated total homocysteine levels reflect the metabolic consequences of methyl donor deficiencies or supplements
2 2010-comparison of the functional properties of pea, chickpea and lentil pr...Bảo Dung Phan
This document compares the functional properties of protein concentrates extracted from pea, chickpea, and lentil using two different processing techniques: isoelectric precipitation (IEP) and ultrafiltration (UF). Optimal extraction conditions were determined for each pulse and variety. Protein content, yield, and functional properties including solubility, water holding capacity, emulsifying properties, foam expansion, and gelling ability were evaluated for the concentrates. Results showed that protein content and functional properties varied depending on the pulse type and extraction method. Red and green lentil concentrates generally had the highest solubility, while yellow pea IEP concentrates had the highest water holding capacity. Chickpea concentrates exhibited better emulsifying
Natural variation in folate levels among tomato (Solanum lycopersicum) Access...Kamal Tyagi
This document describes a study that analyzed natural variation in folate (vitamin B9) levels among tomato accessions. Key findings:
1. Folate levels varied significantly among the tomato accessions, ranging from 4 to 60 μg/100g fresh weight by microbiological assay and 14 to 46 μg/100g by LC-MS.
2. The most abundant folate form was 5-methyltetrahydrofolate (5-CH3-THF) in both mature green and red ripe fruits.
3. Despite the wide variation in folate levels, little polymorphism was found among the accessions in genes involved in folate biosynthesis.
4. The results suggest that variation
Growth Pattern, Molecular Identification and Bio molecules Analysis of FOMITO...journal ijrtem
Abstract : Fomitopsis feei, a brown rot fungus is identified tentatively using morphological characteristics and confirmed phylogenetically by 28S rDNA analysis and sequence was submitted in EMBL Nucleotide Sequence Database. Its growth pattern was studied on eight different solid media and found to be good on Malt extract agar medium. Biomolecules such as proteins and lipid were screened qualitatively and estimated quantitatively. Aminoacid analysis by chromatography and fatty acid analysis by FAME were also done and revealed that tryptophan (20.53%), valine (20.51%) and cis-linoleic acid (43.38%) and palmetic acid (17.88%) were in high percentage.
Key words : Fomitopsis feei, growth, molecular identification and biomolecules
Effects of Metformin, Pioglitazone and Aqueous Extract of Delonix Regia on Bl...iosrjce
The effects of Delonix regia extract (d200mg, d300mg, and d400mg), metformin (m8.3mg, m12.5mg
and m16.5mg), pioglitazone (p0.5mg, p0.7mg and p0.9mg) and combined formulation of metformin and extract
(m6.25d150mg) on glycated hemoglobin status in streptozotocin-induced diabetic Albino wistar rats. Diabetic
status of these rats was assessed by estimating fasting blood glucose levels. A total of 150 albino rats were used
for the investigation and were grouped into twelve groups of twelve rats each as follows; Group I: normal
control rats (NCR). Group II: Diabetic control rats (DCR). Group III: Diabetic rats treated with d200mg.
Group IV: Diabetic rats treated with d300mg. Group V: Diabetic rats treated with d400mg. Group VI: Diabetic
rats treated with m8.3mg. Group VII: Diabetic rats treated with m12.5mg. Group VIII: Diabetic rats treated
with m16.5mg. Group IX: Diabetic rats treated with p0.5mg. Group X: Diabetic rats treated with p0.75mg.
Group XI: Diabetic rats treated with p1.0mg. Group XII: Diabetic rats treated with m125d300mg each for male
and female respectively, for a total of 56 days. After every two weeks interval of treatment for eight weeks three
rats from each group were sacrificed and blood sample were collected and analyzed for various parameters.
The result obtained showed an elevated level of glycated hemoglobin in diabetic-induced wistar albino rats
compared with normal control rats. However, there was reversal of the effects when treated with the
drug/extract. Also there was reduction in the blood glucose level of the diabetic rats treated with metformin
(from 6.37±0.69 to 5.20±0.62mmol/l), pioglitazone (from 7.30±0.21mmol/l to 4.70±0.46), aqueous extract of
Delonixregia (from 8.20±0.81mmol/l to 6.10±0.60) and combined formulation of metformin and extract (from
7.81±0.34 to 4.80±0.17), at p<0.05 confidence level when compared with diabetic control rats in the various
weeks of treatment respectively
This study was designed to evaluate the effect of
70% ethanolic crude extract of Portulaca oleracea L on mice
orgons . (In vivo),In vivo, the acute toxicity of 70 % ethanolic
extract of the plant on normal mice was studied. No toxic effect
was noted on normal mice even at 9500 mg /kg B.W S/C
injection.Histopathological changes due to ethanolic extract of
the plant in healthy mice were summarized in hyperplasia of
white pulp with amyloid deposition, proliferation of
megakaryocytes and mononuclear cell infiltration in the liver and
kidney parenchyma. There were no significant lesions detected in
the brain, heart and ovary in all treated groups.
This document discusses using metabolomics to analyze the food metabolome - all metabolites derived from digestion and metabolism of food components. It notes the complexity of nutritional exposures from foods, which contain nutrients, non-nutrients from natural and man-made sources, and contaminants. Individual metabolic capacity is also influenced by genetics, microbiota, age, and other factors. The food metabolome is defined and applications are described, including discovering new food bioactives, understanding diet-health relationships, and personalized nutrition. Challenges in analyzing the large and diverse food metabolome are also outlined. Initial studies using targeted and untargeted metabolomics are discovering new biomarkers of food intake from various foods. Larger cohort studies and controlled interventions are needed
This study evaluated changes in bioactive compounds and antioxidant activity in the pulp and seeds of Bunchosia glandulifera fruit during different ripening stages. Key findings:
1) Levels of phenolic compounds, carotenoids, vitamin C, and caffeine increased from the second ripening stage to the ripe fruit stage.
2) Ripe fruit had the highest levels of bioactive compounds compared to earlier stages.
3) Phenolic compound and carotenoid levels were highest in the pulp.
4) Carotenoid content was positively correlated with antioxidant activity.
Biochemical effect of Aqueous Carica papaya Seed and Leaf Extracts on Serum B...iosrjce
The cost of administering modern anti diabetic drugs is usually beyond the reach of most people in
the low income group especially those in the developing world where the disease is on the increase. This has led
to the current shift to the use of preparations from various parts of plants because of the current increase in the
knowledge of their toxicity, side effects, active constituents and doses. This study was conducted to compare the
effects of the aqueous Carica papaya seed and leaf extracts on serum biochemistry of alloxan induced diabetic
rats especially glucose level. Male Wister rats weighing 150-200g were induced with single freshly prepared
alloxan monohydrate (150 mg/kg body weight).Diabetes was confirmed after seven days in alloxan-induced rats
showing fasting blood glucose levels ≥ 200mg/dl. The diabetic rats were randomly allocated into three
experimental groups which received Seed extract, leaf extract or normal saline depending on the group. The
extracts were administered orally for twenty-eight days after which the animals were sacrificed and blood
samples were collected for Biochemical analyses. The results showed that both extracts have significant
hypoglycaemic, hepatoprotective and nephroprotective effects although extract of the seed proved to be more
potent than that of the leaf
This document provides an overview of hepatoprotective activity and agents. It discusses the anatomy and functions of the liver, common liver disorders, and mechanisms of hepatotoxicity. Screening methods for evaluating hepatoprotective effects include in vitro, ex vivo, and in vivo models. Several plants are described that have shown hepatoprotective properties, including Picrorrhiza kurroa, Orthosiphon stamineus, Tridax procumbens, Boerhaavia diffusa, and Hoslundia opposita. The document provides references for further information.
Extraction of Colorant From Dragon Fruit Peel and It's Application in a baker...Salemir Hossain
This is a project or thesis proposal presentation for B.Sc Engineering level on Extraction of Colorant From Dragon Fruit Peel and It's Application in a bakery Product. If any help is needed then contact on salemirhossainjust@gmail.com
Diabetes mellitus was induced in rats using streptozotocin. Creatinine, urea, and inflammatory biomarkers were measured in the rats' blood serum. Creatinine and urea levels were significantly increased in diabetic rats, while an inflammatory marker (IL-10) decreased. Administration of Jatropha curcas extracts to diabetic rats resulted in reductions in creatinine, urea, and inflammatory biomarkers, similar to effects of the antidiabetic drug glibenclamide. The extracts showed protective effects against kidney disorders associated with diabetes mellitus.
The document summarizes a study that analyzed samples collected from diarrhea patients and their mothers in rural Bangladesh to detect coliform bacteria. Several species of coliform bacteria were isolated from the samples, with isolation rates ranging from 38.01-3.51%. Extracts of Moringa oleifera leaves were tested against the isolated bacteria. The leaf extracts showed antibacterial effects and were able to inhibit the growth of all tested bacterial pathogens, including E. coli, Shigella, Salmonella, Enterobacter, Klebsiella, and Serratia species. The extracts had zone of inhibition ranging from 8-23 mm and minimum inhibitory concentration values between 62.5-1000 μg/mL, suggesting M. oleifera extracts could
Fruit and Vegetable Waste Hydrolysates as Growth Medium for Higher Biomass an...Premier Publishers
Fruit and vegetable wastes include peels, pulp and seeds that constitute about 40% of the total mass and constitute huge environmental problems. Cultivation of microalgae that utilizes fruit and vegetable wastes as feedstock to produce value added products such as biomass and lipids is a unique approach. Different concentrations of fruit waste hydrolysate (FWH) and vegetable waste hydrolysate (VWH) were used for heterotropic cultivation of Chlorella vulgaris thereby optimizing the suitable hydrolysate concentration for higher biomass and lipid production. FWH in the ratio of 8:2 has produced maximum specific growth rate of 1.92 µ d-1. Higher biomass was recorded in growth medium supplemented with FWH (0.16 mg L-1) than VWH medium. Highest chlorophyll content of 7.2 mg L-1 was observed in 8:2 ratio of FWH whereas it was 4.3 mg L-1 in VWH at the same concentration. Carotenoid content was highest in VWH than FWH media with a maximum content of 0.52 and 0.42 mg L-1 respectively. Fruit waste hydrolysates significantly increased the total lipid content than the vegetable waste hydrolysate medium. Highest lipid content of 6.63 mg L-1 was recorded in 8:2 ratio of FWH. This work demonstrates the feasibility of fruit waste hydrolysate as a nutrient source for algal cultivation and a cost reduction of growth medium in algal biomass and lipid production.
Proximate and Toxicological Analyses of Detoxified Jatropha Curcas Seedsiosrjce
The need for detoxification of less utilized crops with similar nutritional composition with soybeans
as potential substitute or supplement for soybeans in conventional feeds production is obvious as price of
soybean continue to rise. Jatropha curcas is one of such crops which can serve as a potential source of dietary
energy and protein. However, the presence of anti-nutritional factors restricts the utilization of the Jatropha
curcas seed in animal feed. Several researchers however have shown that this obstacle can be overcome by
detoxifying the seeds, but many of them failed to established the effects of these detoxification methods on the
nutritional content of Jatropha curcas .The main objective of this study therefore was to determine the effect of
three simple inexpensive physical methods of detoxification (soaking, roasting and fermentation) on the
proximate and toxicological compositions of Jatropha curcas seed meal. To achieve this, Jatropha curcas seeds
sample used were divided into four parts. The first three parts were subjected to the three different physical
treatments after which they were dried to constant weight and while the fourth part was dried to constant weight
and milled. These four samples were then analysed for their proximate and toxicological composition .The
results showed that fermentation deactivated the antinutrients most in the seeds and did not adversely affect the
nutritional composition of the seeds
This research article evaluated the potential aphrodisiac effects of Moringa oleifera seed extracts in male albino rats. Researchers administered aqueous, alcohol, and chloroform extracts of M. oleifera seeds to male rats for 21 days. They observed several parameters of sexual behavior and found that the extracts significantly increased mounting frequency, intromission frequency, and ejaculation latency, while decreasing mounting latency, intromission latency, and post-ejaculatory interval. The extracts also increased libido without any adverse effects. The results suggest that M. oleifera seed extracts can enhance sexual behavior in male rats, supporting its traditional use as an aphrodisiac.
The International Journal of Engineering and Science (The IJES)theijes
The International Journal of Engineering & Science is aimed at providing a platform for researchers, engineers, scientists, or educators to publish their original research results, to exchange new ideas, to disseminate information in innovative designs, engineering experiences and technological skills. It is also the Journal's objective to promote engineering and technology education. All papers submitted to the Journal will be blind peer-reviewed. Only original articles will be published.
Phytochemical, Proximate and Nutrient Analysis of Cassia Tora Seedsinventionjournals
Recently Proximate analysis and phytochemical analysis and nutrient analysis of seeds of Cassia tora L. from the campus of Government Vidharbha Institute of Science and Humanity, Amravati had been investigated. The seeds sample contained tannin, saponin, protein, steriods, terpenoids, carbohydrate, alkaloids, flavonoids and glycosides. Proximate analysis of moisture, ash, fat amd mineral analysis of calcium, magnesium, iron, nitrogen and solubility were check. The values of it is moisture (56%), cold water (52%), hot water (54%), 1%NaOH (43%), 1%HCl (61%), benzene +alcohol (35%), ash content (17%). These results indicate that the seeds of these Cassia tora L. contains mineral and nutrients elements that will be useful in nutrition. Also the existence of some phytochemicals like tannin, saponin and steroids illustrated medicinal action of the plant in its therapeutic uses. The result of their phytochemcial screening could justify the observed activities and validate their use in herbal medicine.
This project report summarizes research conducted at the Central Science Laboratory (CSL) in the UK on assessing genetic variability and purifying bioactive proteins from oilseed rape (OSR) meal. The research was jointly funded by the European Commission and Home Grown Cereals Authority (HGCA) as part of a larger EU project involving 14 partners.
The studies at CSL showed that DNA fingerprinting, particularly microsatellite analysis, could be used to estimate genetic variability in protein composition among OSR varieties. Research also involved isolating and characterizing antifungal and proteinase inhibitor proteins from OSR meal that have potential applications in plant protection.
The overall EU project aimed to explore value-added uses for OS
Protective Effect of Leaves of Ficus carica Against Carbon Tetrachloride-Indu...UKJPB Journal
The document summarizes a study that evaluated the hepatoprotective effects of leaves of Ficus carica (fig leaves) against carbon tetrachloride (CCl4)-induced liver damage in rats. Various extracts (petroleum ether, ethyl acetate, methanol) of F. carica leaves were prepared and screened for phytochemicals. The ethyl acetate extract was evaluated for its ability to reduce CCl4-induced increases in serum markers of liver damage (SGOT, SGPT, bilirubin) and protect against histological liver damage. Results showed the ethyl acetate extract significantly reduced the elevated serum markers and reversed histological liver changes caused by CCl4, demonstrating hepatoprotective effects
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
All manuscripts are subject to rapid peer review. Those of high quality (not previously published and not under consideration for publication in another journal) will be published without delay.
Thyroid hormone (T3 and T4) deficiency on the metabolism of vitamin Aiosrphr_editor
Both cabbage and thiouracil exert high impact on thyroid hormone. In the present study it is found that decreasing amount of thyroid hormone (T3&T4) inhibit the metabolism of vitamin A in the freshwater fish Ctepharyngodon idella. The goal of this study was to determine the effect of deficiency of thyroid hormone on the retinoid reserve of the fish due to the administration of cabbage and thiouracil as feed. For this experiment fish Ctepharyngodon idella was kept in different feeding states. First was kept with aquatic weed hydrilla, mass grass, filamentous algae. Second group was kept with chopped cabbage and third group was feed with chopped cabbage along with thiouracil. Blood samples were collected from the caudal vasculature in the heparin zed syringes and plasma was assayed for plasma T3&T4 concentration by Radioimmunoassay(RIA). The vitamin A concentration i.e., retinol, dehydroretinol and β-carotene are estimated from the liver of the fish through HPLC procedure. The result shows concentration of mean plasma thyroid hormone was significantly decreased in fishes subjected to cabbage and thiouracil in their feed compared to those of fishes in controlled condition. It is also seen that retinol, dehydroretinol and β-carotene concentration also decreased to a considerable amount in fishes subjected to cabbage and thiouracil in their feed. So, from the present experiment, it is clear that thyroid deficiency can affect the retinoids reserves of fish.
1. Methyl donors such as folate, betaine, choline, and methionine are important nutrients that provide methyl groups for critical cellular processes like DNA methylation and protein synthesis.
2. Betaine donates methyl groups to homocysteine via the betaine homocysteine methyltransferase (BHMT) pathway in the liver and kidneys, the major organs involved in methyl group metabolism. This pathway is regulated by factors like S-adenosylmethionine.
3. Methyl donor deficiencies can disturb energy metabolism and protein synthesis in the liver and cause fatty liver or muscle disorders. Elevated total homocysteine levels reflect the metabolic consequences of methyl donor deficiencies or supplements
2 2010-comparison of the functional properties of pea, chickpea and lentil pr...Bảo Dung Phan
This document compares the functional properties of protein concentrates extracted from pea, chickpea, and lentil using two different processing techniques: isoelectric precipitation (IEP) and ultrafiltration (UF). Optimal extraction conditions were determined for each pulse and variety. Protein content, yield, and functional properties including solubility, water holding capacity, emulsifying properties, foam expansion, and gelling ability were evaluated for the concentrates. Results showed that protein content and functional properties varied depending on the pulse type and extraction method. Red and green lentil concentrates generally had the highest solubility, while yellow pea IEP concentrates had the highest water holding capacity. Chickpea concentrates exhibited better emulsifying
2 2010-comparison of the functional properties of pea, chickpea and lentil pr...
Similar to High performance liquid chromatography coupled to mass spectrometry for profiling and quantitative analysis of folate monoglutamate in tomato.PDF
Natural variation in folate levels among tomato (Solanum lycopersicum) Access...Kamal Tyagi
This document describes a study that analyzed natural variation in folate (vitamin B9) levels among tomato accessions. Key findings:
1. Folate levels varied significantly among the tomato accessions, ranging from 4 to 60 μg/100g fresh weight by microbiological assay and 14 to 46 μg/100g by LC-MS.
2. The most abundant folate form was 5-methyltetrahydrofolate (5-CH3-THF) in both mature green and red ripe fruits.
3. Despite the wide variation in folate levels, little polymorphism was found among the accessions in genes involved in folate biosynthesis.
4. The results suggest that variation
Growth Pattern, Molecular Identification and Bio molecules Analysis of FOMITO...journal ijrtem
Abstract : Fomitopsis feei, a brown rot fungus is identified tentatively using morphological characteristics and confirmed phylogenetically by 28S rDNA analysis and sequence was submitted in EMBL Nucleotide Sequence Database. Its growth pattern was studied on eight different solid media and found to be good on Malt extract agar medium. Biomolecules such as proteins and lipid were screened qualitatively and estimated quantitatively. Aminoacid analysis by chromatography and fatty acid analysis by FAME were also done and revealed that tryptophan (20.53%), valine (20.51%) and cis-linoleic acid (43.38%) and palmetic acid (17.88%) were in high percentage.
Key words : Fomitopsis feei, growth, molecular identification and biomolecules
This research article studied the stability of vitamin C in homogenized fruits and vegetables stored at different temperatures. Samples of broccoli, potatoes, spinach, strawberries, oranges, and tomatoes were homogenized and stored under refrigeration (0-5°C), conventional freezing (-10 to -20°C), or ultra-low freezing (<-55°C) for up to 7 days. Vitamin C was most stable in samples stored at ultra-low freezing, with no decrease after 7 days. Under refrigeration, the largest decreases were seen in raw spinach and broccoli, losing an average of 9.5 mg/100g and 33.1 mg/100g respectively after 1 day. With conventional freezing, losses were
In this pdf, you will get information about manufacturing , standardization , validation , processing , etc. for herbal drugs followed by the WHO guidelines.
This document summarizes a research article that evaluated the immunomodulatory potential of crude protein extract from taro (Colocasia esculenta) corms on hematopoietic cells in mice. The crude taro extract stimulated in vitro proliferation of splenocytes from two mouse strains. When administered intraperitoneally to mice, the extract induced splenomegaly and proliferation of total spleen and bone marrow cells. It also promoted in vivo proliferation of B220+ splenocytes and affected levels of mature and immature B cells in bone marrow. The extract represents a source of immunostimulatory proteins with potential applications as food or pharmaceutical additives.
This document compares the vitamin C content of various citrus fruits as measured by two different methods: titration and high-performance liquid chromatography (HPLC). The study found that oranges contain the highest amount of vitamin C according to both methods. While the absolute vitamin C values differed between the two methods, they produced the same order of vitamin C content across the six citrus fruits tested. Both titration and HPLC are suitable for determining vitamin C, but HPLC is considered more accurate and precise due to the potential for interference in the titration method.
Presentation given by Catarina Duarte from the Chemistry and Biology Technological Institute (New University of Lisbon) in the framework of the Emergence Forum Barcelona
Biocat organized the Barcelona Emergence Forum (April 10-11th, 2014, Congress Palace, Montjuïc) supported by the TRANSBIO SUDOE, a translational cooperation project dedicated to innovation in life sciences in South-West Europe. The Barcelona Emergence Forum contributed to bringing together Academics, Companies, Investment Entities, Technology Platforms and Technology Transfer Offices from Spain, France and Portugal to set up collaborative projects on Human Health & Agro-food Innovation.
More information at: http://www.b2match.eu/emergenceforum2014
CYP2A6_HPLC_PK_2015 New Simple Method for Coumarin in Liver Cytochrome of RatsWael Ebied
This document describes the development and validation of a new liquid chromatography method for determining coumarin and its 7-hydroxy metabolite as a marker of cytochrome P450 2A6 activity in rats. The method uses a C18 column and isocratic mobile phase to separate coumarin, 7-hydroxycoumarin, and an internal standard. The method was validated and showed good linearity, precision, and accuracy. This validated method was then applied to study the effect of resveratrol, sulforaphane, and thymoquinone on hepatic CYP2A6 activity in spontaneously hypertensive rats.
detection and detoxification of aflatoxinMimranjaved1
This document provides an overview of a study on the detection and detoxification of aflatoxins in wheat crops. The study aims to evaluate which wheat varieties have lower aflatoxin contamination and which varieties are more vulnerable. It also aims to develop cheap and effective methods for detoxifying aflatoxins. The methodology will involve collecting wheat samples, quantifying aflatoxins using ELISA kits, and testing physical, chemical and biological detoxification methods. The expected outcome is a comparative profile of aflatoxin levels in different wheat varieties and an effective detoxification approach.
This research article studied the anti-obesity effects of Moringa oleifera leaf extract (MEMOL) in rats with high fat diet-induced obesity. Rats were fed a high fat diet to induce obesity over 49 days. Treatment with MEMOL for 49 days significantly reduced body weight, total cholesterol, triglycerides, and LDL levels in obese rats, and increased body temperature, compared to untreated obese rats. MEMOL treated rats also showed decreased levels of liver enzymes and blood glucose. The results indicate that MEMOL attenuated body weight gain in obese rats without affecting food intake, and demonstrated hypoglycemic and hypolipidemic effects, suggesting it may help treat obesity and related disorders.
This research article studied the anti-obesity effects of Moringa oleifera leaf extract (MEMOL) in rats fed a high-fat diet. Rats fed the high-fat diet for 49 days became obese, with increased body weight, total cholesterol, triglycerides, and lowered HDL levels. Treatment with MEMOL at doses of 200 mg/kg and 400 mg/kg for 49 days significantly reduced body weight gain, total cholesterol, triglycerides, and LDL levels in obese rats compared to those fed just the high-fat diet. MEMOL treatment also increased body temperature and lowered liver enzymes, organ weights, and blood glucose levels in obese rats. The results suggest that MEMOL has anti-ob
The document characterizes dragon fruit and provides information on its nutritional composition and health benefits. It discusses the different varieties of dragon fruit, including their physical and chemical properties. It also describes the antioxidant and fiber content of dragon fruit and their functional roles in reducing risks of chronic diseases. The document summarizes research analyzing the antioxidant activity, nutritional profile, and fatty acid composition of red pitaya fruit from Brazil. It finds that the peel has higher antioxidant levels than the pulp and concludes that the peel should not be discarded due to its nutritional and bioactive compound content.
The document analyzes the nutritional composition of Moringa oleifera leaves from South Africa. It finds the leaves contain high levels of protein (30.3%), various minerals including calcium (3.65%), phosphorus (0.3%), and micronutrients like iron (490 mg/kg). Seventeen fatty acids were identified with alpha-linolenic acid being highest (44.57%). Vitamin E content was also high at 77 mg/100g. Overall, the amino acids, fatty acids, minerals, and vitamin profiles indicate Moringa leaves can provide a desirable nutritional balance.
This study measured levels of environmental contaminants in urine samples from 249 Israeli adults. It found widespread exposure to bisphenol A and organophosphate pesticides. Higher bisphenol A levels were associated with Jewish ethnicity, active smoking, secondhand smoke exposure, and frequent microwave use. Organophosphate pesticide exposure was predicted by age, income, and high fruit consumption. The results suggest intake of fruits is an important source of organophosphate exposure for Israelis. The study provides baseline data that can track changes in exposure from policy actions like pesticide restrictions.
Advabced Breeding methods for quality improvement in solanceous vegetables.docxPallavi Wani
Background/Introduction
World Population during 2020 is 7.8 billion, is growing steadily, and it is predicted to reach 9.1 billion by the year 2050. (united nation population division). Per capita calorie consumption of Fruit & Vegetable is expected to more than triple by the year 2050 in south Asia as compared to 2010. To feed growing population FAO projects the need to increase agricultural output by at least 60 per cent in the next decade. Over the coming decades a changing climate, growing World Population, rising food prices, and environmental stresses will have significant yet impact on food security. To fed growing population, FAO projects the need to increase agricultural output by at least 60 per cent in the next decade. India is ranked 94th position out of 104 countries. And it is also leading producer of many vegetables, but it has estimated around 15 per cent population is under nourished and lack of inadequate food intake both in quality and quantity. So thus improvement of nutritive value of crop is the major gole of breeding programme.
By use of conventional breeding method to get the product will take more number of years and labours, so if we start by modern breeding technologies like, Marker assisted selection, Molecular Breeding, Antisense RNA, and CRIPSR/cas-9 etc., along with conventional method of breeding we can save the lot of time to develop the product.
Vegetable crops are rich in dietary micronutrients, antioxidants and functional phytochemicals. Among vegetables solanaceous vegetables are the good source of health- protective dietary elements such as vitamins, minerals and antioxidant in the human diet. These solanaceous family includes Tomato, Brinjal, Chilli and Potato. (Kumari et al., 2017)
Classification of Quality traits
Any degree of excellence is called quality. There are mainly 5 categories-
1. Morphological traits- Related to product appearance which includes fruit size, shape, color and thickness
2. Orgenoleptic traits- Concerned with palatability of the produce, examples are taste, aroma, flavor etc.,
3. Nutritional traits (Neutraceuticals)- These are determine value of the product such as Beta carotene, ascorbic acid, TSS, Protein , vitamins, minerals
4. Undesirable traits- Includes Solanine content in brinjal and browning in potato
5. Other traits- Includes keeping quality, cooking quality
Advantages of Quality Breeding
It enables grower to get the good product
Helps in consumer acceptability
Makes the product more suitable for processing
Provides Food and health security
Quality traits of Solanaceous vegetables
ADVANCED BREEDING METHODS
E. GENETIC ENGINEERING
F. RNA INTERFERENCE TECHNOLOGY (RNAi)
G. CRISPR CAS9- A genome editor
ACHIEVEMENTS
Transgenic Approach for Bio fortification in vegetables crops
Research Regulation
The square tomato is not only special in terms of shape, but it will also improve your quality of life. Because it contains more than 50% lycopene
FUTURE THRUT
1) The document describes research into using agricultural waste materials like apple pomace and peanut shells to enhance citric acid production through solid state fermentation (SSF) using fungal cultures.
2) Initial screening found that a co-culture of Aspergillus ornatus and Alternaria alternata produced the most citric acid (0.46 mg/mL) when grown on apple pomace.
3) Optimization experiments determined the maximum citric acid yield of 2.644 mg/mL was achieved with the co-culture on apple pomace medium at 50% moisture, pH 5, 25g substrate concentration, and 30°C incubation temperature after 48 hours.
Use of date syrup as alternative carbon source for microbial cultivationPremier Publishers
This document discusses using date syrup and date fruit soaked water as alternative carbon sources for producing biomass of Bacillus megaterium. Maximum biomass production of 2.8 g/l and 4.1 g/l was obtained using 8% date syrup and date fruit soaked water respectively, which was greater than when other carbon sources were used. The optimal medium for high biomass production used 8% date syrup as the carbon source and 0.5 g/L (NH4)2SO4 as the nitrogen source, with fermentation at 30°C for 48 hours. Date syrup and soaked water can be used inexpensively to produce biomass through batch fermentations with B. megaterium.
Application of MALDI-TOF Mass Spectrometry for the Assessment of Prevalence o...Dr. Rahul Suryawanshi
Background: Foodborne infections like Listeriosis cover several disorders and are a worldwide public health emergency. Listeria monocytogenes has been isolated from various foodstuffs, including milk and fishes. MALDI-TOF mass spectrometry technique is known for its rapid and accurate identification of bacterial organisms.
Methods: In the current research, a total of 360 samples comprising raw milk (130), milk products (125) and freshwater fishes (105) were screened for the detection of pathogenic Listeria species by using the USDA method. The recovered Listeria isolates were characterized using conventional set of biochemical analysis along with sugar fermentation tests and further confirmed by MALDI-TOF MS. The virulent nature of pathogenic Listeria isolates was also assessed by in vitro tests like hemolysis on blood agar, CAMP and PI-PLC assay.
Result: In current study, on screening 360 animal origin food samples, three isolates were recovered from raw milk samples and identified as Listeria monocytogenes indicating an overall prevalence of Listeria monocytogenes to the tune of 0.83%. Excellent correlation was observed with identification of Listeria species using conventional phenotypic tests and advanced molecular tool Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) technique. The results depict dependability of advanced technique for rapid and reliable identification of Listeria species.
1) The study investigated the polyphenolic content of rose hip (Rosa canina L.) tea extracts obtained using ultrasound-assisted extraction (UAE) and Soxhlet extraction with different solvents.
2) The highest extract yield was obtained using UAE with water at 619.37 mg/g dried matter. The highest total phenolic content was obtained using Soxhlet extraction with a 50% methanol mixture at 59.69 mg gallic acid equivalent/g dried matter.
3) UAE and Soxhlet extraction with various solvents, including water, ethanol, methanol, and mixtures, were tested on three commercial brands of rose hip tea. Total phenolic content and extract yields
Uchuva: una de las frutas exóticas con grandes propiedades nutricionalesProColombia
This document summarizes a study that used high-performance chemical isotope-labelling LC-MS to identify discriminant metabolites in the plasma of 18 healthy adults after acute and medium-term intake of golden berry fruit. 58 and 43 discriminant metabolites were identified, respectively, after acute and medium-term intake. The identified biological networks mainly involved insulin, epidermal growth factor receptor, and the phosphatidylinositol 3-kinase pathway, showing that berry intake may be associated with insulin signaling and reduce risk factors for metabolic syndrome.
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Gravitropic Analysis of Tomato Seedlings using Time Lapse Video ImagingKamal Tyagi
1. This document describes a protocol for analyzing gravitropism in tomato seedlings using time lapse video imaging. Tomato seeds are sterilized and grown in petri dishes under blue light or infrared light. The plates are rotated 90 degrees to stimulate gravitropic response. Images are captured every 10 minutes for several hours using a webcam.
2. Hypocotyl and root bending angles are measured from the images using image analysis software. Within an hour of reorientation, hypocotyls regain vertical growth orientation in response to gravity. The time course of hypocotyl bending shows the typical sigmoidal gravitropic response curve.
3. This protocol allows real-time observation and quantitative analysis of gravitropism
This document describes a method for rapidly detecting whether plant mutants are homozygous or heterozygous using CEL-I endonuclease. CEL-I endonuclease is a mismatch-specific endonuclease that can cleave DNA at sites of mismatches between DNA strands. The method involves isolating genomic DNA from mutant plants, amplifying a region by PCR, and treating the PCR products with CEL-I endonuclease. Digestion products are then analyzed by gel electrophoresis. For homozygous mutants, no cleavage will occur as there are no mismatches, while heterozygous mutants will show cleavage fragments indicating mismatches. This provides a low-cost method to determine mutational status without sequencing or genetic analysis.
tomato fruit show wide phenotypic diversity but fruit developmental gene show...Kamal Tyagi
- Researchers analyzed 127 tomato accessions from 20 countries and found extensive diversity in fruit traits, clustering them into 9 classes based on traits like soluble solids, carotenoids, ripening index, weight, and shape.
- They screened 10 genes involved in plant development for SNPs using EcoTILLING and found 36 non-synonymous and 18 synonymous changes, identifying 28 haplotypes. However, genetic diversity in the fruit development genes was low compared to the wide phenotypic diversity observed.
- While fruit shape was found to be a complex trait influenced by multiple factors, 100% variance between round and flat fruits was explained by one discriminant function. The study indicates potential to further exploit the morphological and genetic diversity in the
This document describes a method for rapidly detecting whether plant mutants are homozygous or heterozygous using CEL-I endonuclease. CEL-I endonuclease is a mismatch-specific endonuclease that can cleave DNA at sites of mismatches between DNA strands. The method involves isolating genomic DNA from mutant plants, amplifying a region by PCR, and treating the PCR products with CEL-I endonuclease. Digestion products are then analyzed by gel electrophoresis. For homozygous mutants, no cleavage will occur as there are no mismatches. For heterozygous mutants, cleavage will occur at mismatch sites, producing distinctive banding patterns that allow identification of heterozygotes. This method provides a low
Gravitropic Analysis of Tomato Seedlings using Time Lapse Video ImagingKamal Tyagi
1. This document describes a protocol for analyzing gravitropism in tomato seedlings using time lapse video imaging. Tomato seeds are sterilized and grown in petri dishes under blue light or infrared light to observe the gravitropic response without interference from phototropism. Seedlings are imaged every 10 minutes as they bend in response to reorientation of the gravity vector. Hypocotyl bending angles are measured over time to analyze the kinetics of the gravitropic response. The protocol allows real-time observation and quantitative analysis of gravitropism in tomato seedlings.
High performance liquid chromatography coupled to mass spectrometry for profi...Kamal Tyagi
This document describes a study that developed a high performance liquid chromatography coupled to mass spectrometry (HPLC-MS/MS) method for profiling and quantifying folate monoglutamates in tomato fruits. Several parameters of the folate extraction process from tomato tissues were optimized, including extraction conditions, pH range, amount of enzymes, and boiling time. The method was then validated for linearity, sensitivity and recovery. This HPLC-MS/MS method was successfully applied to quantify folates in other plants like spinach, capsicum, and garden pea, demonstrating its versatility for accurate determination of different folate monoglutamates in vegetables.
High performance liquid chromatography coupled to mass spectrometry for profi...
High performance liquid chromatography coupled to mass spectrometry for profiling and quantitative analysis of folate monoglutamate in tomato.PDF
1. High performance liquid chromatography coupled to mass spectrometry
for profiling and quantitative analysis of folate monoglutamates
in tomato
Kamal Tyagi, Pallawi Upadhyaya, Supriya Sarma, Vajir Tamboli, Yellamaraju Sreelakshmi,
Rameshwar Sharma ⇑
Repository of Tomato Genomics Resources, Department of Plant Sciences, School of Life Sciences, University of Hyderabad, Hyderabad 500046, India
a r t i c l e i n f o
Article history:
Received 21 October 2014
Received in revised form 20 January 2015
Accepted 23 January 2015
Available online 31 January 2015
Keywords:
Tomato
Folates
HPLC–MS/MS
Folate extraction
a b s t r a c t
Folates are essential micronutrients for animals as they play a major role in one carbon metabolism.
Animals are unable to synthesize folates and obtain them from plant derived food. In the present study,
a high performance liquid chromatography coupled to mass spectrometric (HPLC–MS/MS) method was
developed for the high throughput screening and quantitative analysis of folate monoglutamates in
tomato fruits. For folate extraction, several parameters were optimized including extraction conditions,
pH range, amount of tri-enzyme and boiling time. After processing the extract was purified using
ultra-filtration with 10 kDa membrane filter. The ultra-filtered extract was chromatographed on a RP
Luna C18 column using gradient elution program. The method was validated by determining linearity,
sensitivity and recovery. This method was successfully applied to folate estimation in spinach, capsicum,
and garden pea and demonstrated that this method offers a versatile approach for accurate and fast
determination of different folate monoglutamates in vegetables.
Ó 2015 Elsevier Ltd. All rights reserved.
1. Introduction
Folate (folic acid) is one of the B groups of vitamins that are
essential for the human health. The dietary insufficiency of folate
can lead to several health related disorders such as megaloblastic
anemia (Gough, Read, McCarthy, & Waters, 1963), exacerbation
of cardiovascular disease (Kolb & Petrie, 2013) and some types of
cancer (Blount et al., 1997). It is also required for the development
of a healthy fetus and its deficiency affects formation of fetus’
spinal cord and brain (Blom, Shaw, den Heijer, & Finnell, 2006).
Folate cannot be synthesized by animals and therefore it is solely
obtained from diet.
Plants are the major source of dietary folates. Green leafy vege-
tables, legumes and some fruits are among the richest sources of
folate. In several countries the cereal based food products are
mandatorily fortified with folic acid to prevent folate-related disor-
ders. In addition, transgenic approaches have been used to increase
the biosynthesis of folates in tomato fruits, potato tubers and rice
grains to remove dietary constraints of low folate levels in diet
(Blancquaert, De Steur, Gellynck, & Van Der Straeten, 2014).
Folate is comprised of a pterin moiety attached by a methylene
bridge to para-amino benzoic acid, which is coupled to one or more
glutamyl residues. In vivo folates exist as tetrahydrofolate (THF)
and its derivatives (5-methyl, methylene, methenyl, or 10-formyl)
which vary in oxidation states, single carbon substituents, and
with a variable number of glutamyl residues (Rébeillé et al.,
2006), which are collectively called – folate or vitamin B9. THF
plays a key role in one-carbon transfer reactions in all living organ-
isms participating in diverse metabolic reactions such as amino
acid metabolism, pantothenate synthesis, purines and thymidylate
synthesis etc.
For quantitative determination of total folates, microbiological
assay is the most commonly used method which is also recom-
mended by Association of Official Analytical Communities Interna-
tional (AOAC, 2000). The main weakness of this method is that it
cannot distinguish diverse forms of folate present in food samples.
Above limitation has been overcome by using chromatography-
based methods allowing separation of different vitamers. The
folate determination has been carried out using high-performance
liquid chromatography (HPLC) coupled with UV (Pfeiffer, Rogers, &
Gregory, 1997), electrochemical (Bagley & Selhub, 2000), and
fluorescence detection (Ndaw, Bergaentzlé, Aoudé-Werner,
http://dx.doi.org/10.1016/j.foodchem.2015.01.110
0308-8146/Ó 2015 Elsevier Ltd. All rights reserved.
⇑ Corresponding author. Tel.: +91 40 23010514; fax: +91 40 23010120.
E-mail addresses: tyagi.kamal6672@gmail.com (K. Tyagi), pravas43@gmail.com
(P. Upadhyaya), supu.megha@gmail.com (S. Sarma), vajirchem@gmail.com (V.
Tamboli), syellamaraju@gmail.com (Y. Sreelakshmi), rameshwar.sharma@gmail.
com (R. Sharma).
Food Chemistry 179 (2015) 76–84
Contents lists available at ScienceDirect
Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem
2. Lahély, & Hasselmann, 2001). The folate level in tomatoes was esti-
mated using HPLC coupled with fluorescence detector (Zhang et al.,
2003). However, HPLC is unable to quantify folate forms occurring
in very low amounts.
In recent years, HPLC/UPLC hyphenated with mass spectrometry
has allowed both qualitative profiling and quantification of folate
derivatives in several foods, because of its high sensitivity, selectiv-
ity, specificity, and accuracy (De Brouwer, Zhang, Storozhenko, Van
Der Straeten, & Lambert, 2007; Garratt et al., 2005; Patring &
Jastrebova, 2007; Rychlik & Freisleben, 2002; Zhang, Storozhenko,
Van Der Straeten, & Lambert, 2005). The gas chromatography mass
spectrometry (GC–MS) has also been used for quantitative analysis
of total folates using acidic extraction (Dueker et al., 2000). How-
ever, GC–MS requires derivatization of all folate to para-amino ben-
zoyl glutamate prior to analysis hindering distinction of different
folate forms, thereby limiting its usage.
The precise analysis of folate derivatives present in plants/food
samples is a complicated task stemming from multiplicity of folate
derivatives, bound/unbound forms, low levels of folate and differ-
ences in plant matrix such as seeds, leaf, fruits or tuber. The deter-
mination of folate in food is also strongly influenced by the sample
preparation and extraction methods. Given that each plant matrix
is distinct, there is no standard method for extraction of folate from
plant material owing to difficulties in sample preparation, extrac-
tion, deconjugation and purification. This entails that for each
plant/tissue the extraction protocol has to be independently opti-
mized to enable precise qualitative and quantitative determination
of folate levels (Arcot & Shrestha, 2005). Currently limited methods
are available to extract and specifically quantify different folate
derivatives present in plants, such as in spinach (Zhang et al.,
2005), rice (De Brouwer et al., 2008), and potato (Van Daele
et al., 2014).
The aim of this study was to develop an accurate, reproducible,
and quantitative method for high throughput screening of large
populations of different cultivars, mutants and natural accessions
of tomato. We report here optimization of extraction, validation,
and application of a high performance liquid chromatographic
method with mass spectrometric (HPLC–MS/MS) detection for
quantification of folate monoglutamates in tomato. The all ion
fragmentation approach of the Orbitrap mass spectrometer
allowed confirmation of all targeted folate monoglutamates at
the same time without pre-selection. The above method was also
used for quantification of folates from plants such as capsicum
fruits and spinach leaf.
2. Materials and methods
2.1. Chemicals and reagents
5-Methyltetrahydrofolate (5-CH3-THF), tetrahydrofolate (THF),
5,10-methenyltetrahydrofolate (5,10-CH+
THF), 5-formyltetrahy-
drofolate (5-CHO-THF) and 5,10-methylenetetrahydrofolate
(5,10-CH2-THF) were purchased from Schirck’s Laboratory (Jona,
Switzerland). Folic acid (FA) and methotrexate (MTX) were pur-
chased from Sigma–Aldrich Co. (St. Louis, USA). Supplementary
Fig. 1 shows the structure of various folate forms and MTX.
LC–MS grade acetonitrile was purchased from Sigma–Aldrich
Co. (St. Louis, USA). Ultrapure water (18.2 mX at 25 °C) was
obtained from Milli-Q water purification system (Millipore, Brad-
ford, USA). Ascorbic acid and 2-mercaptoethanol (ME) were pur-
chased from Sigma–Aldrich Co. (St. Louis, USA). Formic acid
(HCOOH) and acetic acid (CH3COOH) of LC–MS grade were
purchased from Fisher Scientific (Loughborough, UK). Potassium
dihydrogen phosphate, dipotassium hydrogen phosphate and acti-
vated charcoal were purchased from HiMedia (Mumbai, India).
2.2. Preparation of standard and enzyme solutions
Stock solutions of folate standards (1 mg/mL) were prepared in
50 mM potassium phosphate solution, pH 4.5 containing 1% (w/v)
of ascorbic acid and 0.5% (w/v) of 2-mercaptoethanol except FA
and MTX, which were dissolved in neutral or basic pH buffer.
The standard stock solutions were diluted appropriately in the
extraction buffer to prepare working solutions. The remaining
stock solutions were flushed with nitrogen gas, and small aliquots
were stored at À80 °C.
Protease (from Streptomyces griseus, RM6186) and a-amylase
(from Bacillus sp., A6814) were purchased from HiMedia (Mumbai,
India) and Sigma–Aldrich Co. (St. Louis, USA) respectively. Protease
(2 mg/mL) and a-amylase (20 mg/mL) were dissolved in ultrapure
water, and aliquots were stored at À20 °C. Rat plasma was pur-
chased from National Institute of Nutrition (NIN), Hyderabad,
India. To remove endogenous folate from rat plasma, 100 mL of
rat plasma and a-amylase were mixed with 5 g of activated char-
coal separately, stirred on ice for 1 h followed by centrifugation
at 5000g for 10 min at room temperature. The supernatant was fil-
tered through a 0.22 lm filter, divided into 1 mL aliquots, and
stored at À20 °C. Protease was used without pre-treatment and
was stored at À20 °C. For ultra-filtration, 10 kDa membrane filters
(Pall Corporation, USA) were used.
2.3. Plant material: growth conditions and tissue harvesting
Tomato plants (Solanum lycopersicum L. cv. Arka Vikas) were
grown in a greenhouse of University of Hyderabad at 25 ± 2 °C
day and ambient temperature in night. For extraction procedure,
method development, and validation experiments, different tissues
were selected including tomato fruit and leaf tissue, garden pea
seeds, capsicum fruit and spinach leaf. Fresh garden pea, spinach
and capsicum were purchased from the local market. The juvenile
leaves from six-weeks-old plants and fruit tissue at mature green
and red ripe stage were harvested from tomato plants. Tomato
fruits, pea seeds, and capsicum fruits were homogenized to powder
after freezing in liquid nitrogen using a homogenizer (IKA A11,
Germany). The tomato and spinach leaves were manually homog-
enized in liquid nitrogen using pre-chilled mortar and pestle.
Homogenized powder was used for extraction of folates.
2.4. Sample extraction: homogenization, enzyme treatments and
purification
The basic extraction procedure for folates was adopted from De
Brouwer et al. (2008) with modifications. Several parameters were
evaluated to obtain the best recovery of folates from tomato fruit
and leaf. These modifications included selection of appropriate
pH and concentration of extraction buffer, amount of enzyme
and different boiling treatment to deactivate enzyme activity.
The extraction buffer (pH 4.5) consisting of 50 mM potassium
phosphate, 1% (w/v) ascorbic acid, 0.5% (v/v) b-mercaptoethanol
and 1 mM calcium chloride was freshly prepared for the sample
extraction and flushed with nitrogen for 20 s. Initially, 650 lL of
extraction buffer was mixed with 100 mg of plant homogenate in
2 mL Eppendorf tube. To the extract 2.7 ng/mL of individual folates
were added at the beginning of extraction process for recovery
calculation.
The tubes were capped tightly and after mixing by vortexing
were placed in a boiling water bath for 10 min. After cooling on
ice, the extract was incubated with 10 lL of a-amylase at
room temperature for 10 min to digest complex carbohydrates.
Thereafter, 2.5 lL of protease was added and tubes were incubated
at 37 °C for 1 h, then transferred to boiling water bath for 5 min.
After cooling on ice, the deconjugation of folate polyglutamates
K. Tyagi et al. / Food Chemistry 179 (2015) 76–84 77
3. to monoglutamates was carried out by adding 100 lL of rat plasma
to tubes and incubating at 37 °C for 2 h. Thereafter the sample was
boiled for 5 min followed by cooling on ice. The tubes were centri-
fuged for 30 min (14,000g, 4 °C) and the supernatant was filtered
through the 0.22 lm filter. Finally, the filtrate was clarified by
ultra-filtration at 12,000g for 12 min using 10 kDa cut-off mem-
brane filter. Immediately, the resulting extract was transferred to
autosampler vials in HPLC and a 7.5 lL aliquot was directly
injected on the column.
2.5. Liquid chromatography settings
The chromatographic separation of folate derivatives was car-
ried out using Waters Acquity™ UPLC system (Milford, USA) run-
ning in HPLC mode, coupled to a binary pump, an autosampler,
and controlled by Xcalibur 3.0 software (Thermo Fisher Scientific,
San Jose, USA). The sample manager was configured with a 10 lL
sample injection loop and a 100 lL sample syringe with the partial
loop with needle overfill function. Prior to sample injections, the
syringe was rinsed with 500 lL of weak (water/acetonitrile, 95:5)
and strong (water/acetonitrile, 5:95) solvents, respectively. The
auto-sampler (set at 4 °C) was covered to minimize any degrada-
tion of light sensitive folates during sample analysis.
Folate derivatives were separated on a reversed phase Luna C18
column (5 lm particle size, 250 mm  4.60 mm ID) (Phenomenex,
USA) using gradient elution program. The gradient comprised of a
binary solvent system consisting 0.1% (v/v) of formic acid in water
(solvent A) and acetonitrile (solvent B) at a flow rate of 500 lL
minÀ1
. The injection volume was 7.5 lL, run time 25 min and col-
umn temperature was set at 24 °C. The starting condition of gradi-
ent was 95% of solvent A and 5% of solvent B. Subsequently, solvent
B was linearly increased to 10% in 4 min, then to 30% in 6 min and
to 95% in 7 min. Thereafter, the mobile phase was reverted to the
initial condition in 3 min and held for 5 min for re-equilibration
of column before the next injection. After sample injection for first
8 min, the solvent flow from LC pump was diverted to waste posi-
tion to minimize the suppression of MS signals by salts present in
the extract. Thereafter, the solvent flow was switched to mass
detector position for next 12 min and again diverted to waste posi-
tion for 5 min till completion of the total run.
2.6. Mass spectrometry settings
For all experiments, Exactive™ Plus Orbitrap mass spectrometer
(Thermo Fisher Scientific, USA) was operated in alternating full
scan and all ion fragmentation (AIF) mode equipped with positive
heated electrospray ionization (ESI). The source parameters were
optimized as follows: ion spray voltage, 4 kV; capillary tempera-
ture, 380 °C and heater temperature, 300 °C. Sheath and auxiliary
gas flows were 60 and 20 arbitrary units (au) respectively. Capil-
lary, tube lens, and skimmer voltages were 65, 60, and 24 V respec-
tively. For full scan experiment, the mass scan in the range of m/z
120–800 was monitored, while for AIF experiment, it was in the
range of m/z 50–450. The resolution for full scan experiment was
set to 70,000 full width at half maximum (FWHM) and for the
AIF experiment it was set to 35,000 FWHM. The automatic gain
control (AGC) target for full scan and AIF experiments were set
to high dynamic range of 1e6
and 5e5
respectively. Similarly, the
maximum injection time was set to 100 and 50 ms respectively.
All ions entering into the mass spectrometer (MS) were frag-
mented using nitrogen gas in the High-energy collision-induced
dissociation (HCD) cell at normalized collision energy (CE) 20
(arbitrary unit) with 20% stepped CE, and finally detected in the
Orbitrap mass analyzer. The fore vacuum, high vacuum and
ultra-high vacuum were maintained around 1.5 mbar, from
2.5eÀ5
to 3.5eÀ5
mbar and below 4eÀ10
mbar, respectively.
Instrument control and data processing were carried out by Xcali-
bur 3.0 software (Thermo Fisher Scientific, San Jose, USA).
Prior to sample analysis, the external calibration of instrument
was performed to ensure the mass accuracy by directly infusing
calibration solutions in both positive and negative ion modes.
The positive mode calibration solution consisted of caffeine, Met-
Arg-Phe-Ala acetate salt (MRFA), n-butyl amine and Ultramark™
1621, while negative mode calibration solution consisted of
sodium dodecyl sulfate, sodium taurocholate and Ultramark™
1621. These solutions were freshly prepared using acetonitrile,
methanol, and water containing 1% (v/v) acetic acid. All these cal-
ibration solutions were mixed at room temperature and directly
infused in the source using a syringe pump (Chemyx Fusion 100,
Thermo Fisher Scientific, USA) with a flow rate of 5 lL/min. In posi-
tive ion mode, the external lock mass (polydimethyl cyclosiloxane,
exact mass = 455.12003) was also tested for the mass analyzer
recalibration.
2.7. Recovery studies
In the absence of specific guidelines for analysis of folate deriv-
atives in tomato plant tissue, the International Conference on
Harmonization (ICH) guideline was used for the validation of ana-
lytical method which includes accuracy, sensitivity and linearity.
To determine the accuracy of method, recovery test was performed
by spiking of tomato extract with known amount of individual
folate derivatives and calculating their final content in extract.
The recovery (R) was calculated as B À C/A Â 100, where A = peak
area of neat folates standard, B = peak area of spiked standard,
C = peak area of extract.
2.8. Limit of detection and limit of quantification
Sensitivity was confirmed by evaluating the limit of detection
(LOD; calculated as: 3.3r/S, where r is the standard deviation
and S is the slope of calibration curve) and limit of quantification
(LOQ; calculated as: 10r/S). The linearity of each folate derivative
was evaluated by plotting the peak area at different concentra-
tions. The sample concentrations were calculated from the equa-
tion y = mx + c, as determined by the linear regression analysis.
2.9. Statistical analysis
All results are expressed as mean value ± SE of three or more
replicates based on fresh weight (FW). A t-test was used for deter-
mining significant difference between the mean values. The differ-
ences were considered to be significant for P < 0.05. Statistical
analysis of data was performed using SigmaPlot 11.0.
3. Result and discussion
3.1. Optimization of HPLC and MS/MS conditions
It has been reported that reverse phase C18 column is the most
appropriate column (based on resolution and retention time) for
the separation and detection of folate derivatives. Initially, both
organic solvents (acetonitrile and methanol) with 0.1% (v/v) formic
acid and 0.1% (v/v) acetic acid were evaluated. The results show
that usage of acetonitrile with 0.1% (v/v) formic acid increased
the resolution with higher signal to noise ratio and proved to be
the best eluent for ionization of folate derivatives. The optimized
chromatographic conditions are described in Section 2.5. Under
these conditions, separation of THF, 5-CH3-THF, 5,10-CH+
THF,
MTX, 5-CHO-THF, and FA were achieved at 10.28, 10.84, 11.35,
12.87, 13.02, 13.16 min respectively (Fig. 1). Interfering
78 K. Tyagi et al. / Food Chemistry 179 (2015) 76–84
4. compounds present in the plant extract (sugars, salt, and other
matrices) may suppress ionization. To eliminate them, a divert
valve was used during first 8 min and last 5 min of chromato-
graphic run to redirect the major impurities to the waste instead
to the mass detector.
Prior to determining the optimal conditions, individual folate
standard derivatives and internal standards were directly infused
to ESI in both positive and negative ion modes. Comparison of both
modes revealed that the positive ion mode provided better sensi-
tivity than negative ion mode in conformity with earlier reports
(De Brouwer et al., 2007; Patring & Jastrebova, 2007; Rychlik &
Freisleben, 2002). Therefore, positive ion mode was selected for
subsequent experiments. The optimized MS parameters are
described in Section 2.6. For identification of each compound, accu-
rate mass of the corresponding ion, elemental composition, and
specific retention times were considered. In full scan mode, the
ions detected at m/z 446.1779, 460.1942, 456.1632, 455.1786,
474.1729 and 442.1467 corresponded to the THF, 5-CH3-THF,
5,10-CH+
THF, MTX, 5-CHO-THF and FA respectively. Fig. 1 shows
the extracted ion chromatogram. The accurate masses of corre-
sponding ions were also compared with the theoretical masses cal-
culated by Xcalibur 3.0 software. In addition, all ions were
fragmented without pre-selection in HCD cell at different collision
energy ($20–40 au). Interestingly, several fragment ions were
detected at m/z 299.1249 (THF), 313.1404 (5-CH3-THF), 308.1256
(MTX), 327.1198 (5-CHO-THF), and 295.0936 (FA) indicating the
loss of glutamate moiety (Supplementary Table 1) except
412.1530 (5,10-CH+
THF). Using HCD, the sample analysis time
was substantially reduced by confirming the observed m/z values
at the same time without pre-selection.
3.2. Optimization of sample extraction in tomato
For optimization of folate extraction/estimation from tomato,
several factors were considered like type and composition of sam-
ple matrix, nature, amount, and stability of the compounds for
extraction and fine tuning of the LC–MS parameters for their detec-
tion. Considering that folates are very labile compounds, their
extractions differ for each kind of matrix. Tomato fruit is a very
complex matrix having many compounds to which folates bind.
We optimized the folate extraction from tomato by modifying
the method of De Brouwer et al. (2008) originally developed for
rice. Largely for the folate extraction from a variety of food
matrices, a trienzyme treatment (amylase, protease and conjugase)
developed by Martin, Landen, Soliman, and Eitenmiller (1989) is
used to release the bound folate from matrix. The trienzyme treat-
ment accomplishes more complete extraction of folates that are
bound to matrices of proteins or polysaccharides by including pro-
tease and a-amylase in addition to conjugase.
Considering matrix complexity, several parameters were evalu-
ated for optimal folate extraction from tomato fruits such as pH
and molarity of the extraction buffer, amount of a-amylase, prote-
ase, rat plasma conjugase, and duration of boiling time after
enzyme treatments. Fig. 2 shows optimization of different param-
eters for folate extraction. The pH of extraction buffer greatly
affects the stability of folates in extracts. Therefore, folate extrac-
tion was compared using two different pH, 4.5 and 7.0. No signifi-
cant difference in recovery of total folate content was observed
using these two pH (P value 0.0519) (Fig. 2A).
Stability of the folate derivatives greatly varies with respect to
pH, thus requiring selection of a pH that is optimal for efficient
extraction of most folate derivatives (De Brouwer et al., 2007).
The optimal pH for protease and amylase is also different from
optimal pH for rat plasma conjugase (Arcot & Shrestha, 2005).
Since multiple pH adjustments prolong the extraction process
and increase the chances of oxidation of folate, the usage of differ-
ent pH to achieve optimal trienzyme treatment is not feasible
while screening a large number of samples. The amount of total
folate extracted at pH 4.5 and 7.0 was nearly similar, however, at
pH 4.5, we detected THF in tomato leaf but not in tomato fruit tis-
sue. Given the variability in stability of different folate derivatives,
it is difficult to have an ideal pH for extraction of all folate
derivatives in their native state from plant matrix. Considering that
5-CH3-THF and 5-CHO-THF, the most predominant folate forms
present in the plants are more stable at low pH, therefore we used
pH 4.5 for subsequent extraction of folates. The recovery of folate is
also influenced by the buffer composition; however, phosphate
buffer is most widely used for folate extraction. We compared both
50 and 100 mM phosphate buffer and found that both were equally
efficient. Taking cognizance of fact that high salt concentrations in
buffer interfere with the ionization process in the mass spectrom-
eter, we used 50 mM phosphate buffer for all extraction
procedures.
Considering that the fruit and vegetable matrices are complex,
possessing compounds that may possibly bind with the metabolite
of interest and improve/inhibit the extraction efficiency, the trien-
zyme method of folate extraction was also optimized. The optimi-
zation involved alteration of enzyme concentrations and duration
of treatments. We compared the extraction of folate with and
without respective enzymes for all three enzymes used for folate
Fig. 1. Representative chromatogram of five folates standards and an internal standard. (A) THF, (B) 5-CH3-THF; (C) 5,10-CH+
THF; (D) MTX; (E) 5-CHO-THF and (F) FA.
K. Tyagi et al. / Food Chemistry 179 (2015) 76–84 79
5. extraction. It was observed that without a-amylase treatment,
$85% of the total folate content was extracted (P value 0.0281)
compared to the a-amylase treated control. Similarly, addition of
protease and incubation at 37 °C for 1 h did not significantly
improve the extraction of total folate content (P value 0.050)
(Fig. 2B). Among the three enzymes used in this study, the rat
plasma conjugase was found to be the most important for folate
deglutamylation. Different amounts of rat plasma conjugase was
added to the extract (per 100 mg of fresh tissue), that is, 0, 25,
50 lL which resulted in 17%, 52% and 88% recovery of total folate
pool respectively compared with addition of 100 lL rat plasma
serving as control. Taking into account that increase from 50 to
100 lL of rat plasma lead to 12% improvement in the folate
recovery, 100 lL of rat plasma was used for all subsequent exper-
iments (Fig. 2C).
It is reported that boiling the extract prior to enzyme treatment
reduces the overall yield of folate from food matrices. Moreover,
the addition of antioxidants such as ME and ascorbic acid in the
extraction buffer protects folate from degradation during boiling
treatment that is needed to denature the enzymes besides improv-
ing the stability of folate in an autosampler. The increase in the
boiling time from 5 to 10 min post-enzyme treatment did not
enhance the recovery of folate, therefore, boiling for 5 min was
used for all enzymatic deactivation steps to minimize any possible
degradation of folates (Fig. 2D).
3.3. Optimization of purification
Different folate vitamers differ in their functional groups thus
vary in their chemical properties and therefore can be better sepa-
rated by HPLC. It is reported that sample purification prior to sep-
aration in HPLC leads to better recovery of folate and also
distinction between different forms. The presence of different
endogenous constituents in food extracts interfere with folates
and hamper the determination of some folate forms. Considering
that folate is also present at low level in most food matrices,
removal of the interfering compounds during purification
improves the detection limit and selectivity of the folate in HPLC.
Solid phase extraction (SPE) is the most commonly used method
for folate purification using either a strong anion exchange (SAX)
sorbent or C18 SPE cartridges for food extract prior to HPLC–MS
analysis (Chandra-Hioe, Bucknall, & Arcot, 2013; Stokes & Webb,
1999). Use of SAX sorbent for purification of food extracts provides
high recovery of different folate forms, however, pre-concentration
of sample is not possible as large buffer volumes are required to
elute all the folates from SAX column. In addition, high concentra-
tion of salt used in elution step interferes with subsequent MS
analysis.
Recently, affinity chromatography using immobilized folate-
binding protein has been used as a highly selective purification
method for folates, especially for complex food matrices such as
cereals (Pfeiffer et al., 1997). Though affinity chromatography
enables quantification at 10-fold lower concentration than SAX,
the lack of commercial availability of FBP columns precludes their
use for folate analysis. Moreover, folate-binding protein exhibits
low affinity to 5-HCO-H4 folate, which may result in higher losses
of this folate form during the purification step.
Among the different methods of folate analysis, usage of LC–MS
has emerged as most preferred method owing to its specificity and
sensitivity to distinguish different forms of folate. Several studies
have used LC–MS for folate analysis, particularly from fortified
food samples (Alaburda, de Almeida, Shundo, Ruvieri, & Sabino,
2008; Patring, Wandel, Jägerstad, & Frølich, 2009). However, the
detection of folate derivatives by LC–MS needs robust purification
of folate vitamers for efficient and accurate analysis. In this study,
we optimized the purification of folates for subsequent analysis by
LC–MS. In earlier studies, SPE using SAX cartridge (Vishnumohan,
Arcot, & Pickford, 2011) or affinity column using folate binding
protein (Díaz de La Garza et al., 2004) have been used so far. How-
ever, these approaches are time consuming and cannot be applied
when handling large number of samples. To overcome these draw-
backs which hinder folate estimation for high throughput analysis,
the purification step was modified to a simple, efficient and also
cost effective method for quantification of folate in tomato by
using a molecular weight cut-off membrane filter according to
Zhang et al. (2005). For efficient sample cleanup, extract was fil-
tered through a 0.22 lm filter to remove the particulate matter
in the extract followed by ultra-filtration through a 10 kDa
Fig. 2. Optimization of folate extraction in tomato fruit. (A) Effect of pH, (B) extraction in presence (control) and absence of a-amylase and protease, (C) extraction with
increasing amount of rat plasma conjugase and (D) effect of duration of boiling (n P 3).
80 K. Tyagi et al. / Food Chemistry 179 (2015) 76–84
6. centrifugal device removing various polymeric compounds present
in the tomato extract. The usage of step wise increase in gradient
consisting of 0.1% (v/v) of formic acid in water (solvent A) and ace-
tonitrile (solvent B) on a C18 reverse phase column yielded good
resolution of different folate vitamers. To avoid contamination of
electrospray of MS machine by salt and matrix, which reduces
the sensitivity of the mass spectrometer, out of total runtime of
25 min, first 8 min and last 5 min eluent were discarded to the
waste and sample was allowed to pass through MS during inter-
vening 12 min.
Folates were analyzed in positive ion mode through mass spec-
trometry as the analysis indicated it to be superior to the negative
ion mode. All ion fragmentation (AIF) parameters including precur-
sor ions, product ions, and collision energy were optimized for frag-
mentation by injecting the folate standards. Electrospray ionization
(ESI) source temperature was 300 °C and capillary temperature was
maintained at 380 °C. For fragmentation of the folate standards in
HCD cell, collision energy of 20 or 40 (arbitrary unit, Supplementary
Table 1) was considered optimal depending on the folate com-
pound. The most abundant fragment resulting from all folates
was the one that was generated from the neutral loss of the gluta-
mate moiety (m/z 147), e.g. for FA m/z 442/295, for THF m/z 446/
299, for 5-CH3-THF m/z 460/313, for 5-CHO-THF m/z 474/327 and
for MTX m/z 455/308. Collision energy used for 5,10-CH+
THF was
40 arbitrary unit resulting in m/z 456/412. Supplementary Table 1
shows the fragmentation result of the folate standards. A represen-
tative chromatogram of folates standards is shown in Fig. 1.
3.4. Standard calibration curve and linearity
The linearity of the developed HPLC–MS method was evaluated
by preparing seven point calibration curves without matrix for
each folate derivative (THF; 5-CH3-THF; 5,10-CH+
THF; MTX;
5-CHO-THF; and FA) in triplicates. Correlation coefficients (R2
)
determined for all these folates were approximately 0.99, which
confirmed their good linearity within the considered concentration
ranges. For the folates derivatives, which were detected in the
tomato fruits, the limit of detection (LOD, S/N P 3.3) and quantita-
tion (LOQ, S/N P 10) were theoretically determined based on cali-
bration curve, relating concentrations with signal to noise ratios.
Table 1 shows the correlation coefficient, linear range, and slope
of calibration curve.
3.5. Recovery and stability
Accuracy of the developed method was determined by recovery
test. The mean recoveries (n = 3) for all individual folate derivatives
were in the range of 48–131% (Table 2). The observed value indi-
cates that the developed method is accurate, except for folic acid,
which is not a naturally occurring folate. Taking cognizance that
multiple samples were analyzed for folate levels, the stability of
the extracted samples was examined in the autosampler at 4 °C.
The results indicated that most of the folate derivatives are stable
up to 72 h of storage in the autosampler at 4 °C barring 5,10-CH+-
THF (Supplementary Fig. 2). It is reported that the presence of
ME and ascorbic acid in the extraction buffer enhances the stability
of the folate for a prolonged period (Vahteristo & Finglas, 2000).
3.6. Use of external and internal standard
In order to check the possible variations in MS response for dif-
ferent folate derivatives while screening of large number of sam-
ples, we used internal standards for accurate and reproducible
quantitation. Though the usage of stable isotope labeled folate
standards is the best option, these were not used owing to their
limited availability and high cost. In its place a known amount of
individual folate derivatives were used as external standards. The
MS response of folate derivatives were checked by analyzing sev-
eral tomato extracts and comparing the calculated concentrations
with initial values. Furthermore, we used two structurally similar
compounds, FA and MTX (Zhang et al., 2005) as internal standards.
Both these standards were mixed to the tomato homogenate prior
to initiation of extraction protocol and recoveries were calculated.
Considering that the recovery of MTX was higher than FA (Table 2),
it was selected as internal standard for our further screening.
3.7. Folate content in tomato fruit
The total folate level in mature green and red ripe fruits of
tomato cultivar Arka Vikas was 21.94 and 18.06 lg/100 g of FW
respectively (Fig. 3A). 5-CH3-THF was the most predominant folate
form comprising of 68% and 58% of total folate in mature green and
red ripe fruits respectively. These values were similar to those
reported by Díaz de La Garza, Gregory, and Hanson (2007) that
5-CH3-THF constitutes 70% of total folate present in tomato fruit.
In addition to 5-CH3-THF, other two folate derivatives 5-CHO-
THF and 5,10-CH+
THF were also detected in tomato fruit. Among
these, 5-CHO-THF was the second predominant folate form present
in both mature green and red ripe stage fruits. The level of 5-CH3-
THF significantly decreased during tomato fruit ripening from
mature green to red ripe stage (P value 0.035). In contrast, the con-
tent of 5,10-CH+
THF and 5-CHO-THF in the mature green and red
ripe stage of tomato fruit did not show any significant changes
(Fig. 3B). Considering that 10-CHO-THF cannot be identified in
the LC–MS due to its conversion to 5,10-CH+
THF in mobile phase
containing 0.1% formic acid (Fazili & Pfeiffer, 2004; Goyer et al.,
2005), it can be assumed that 5,10-CH+
THF peak likely included
both 10-CHO-THF and any preexisting 5,10-CH+
THF. Consistent
Table 1
Calibration and sensitivity data of folate standards. All experiments were done in triplicates.
Compounds LOD (ng/mL) LOQ (ng/mL) Slope (n = 6 or 7) mean ± SE R2
Linear range (ng/mL)
FA 0.15 0.47 38095.5 ± 450.5 0.999 1–30
THF 0.25 0.77 30541.5 ± 577.5 0.997 2–30
5,10-CH+
THF 0.34 0.92 10757.3 ± 70.7 0.984 0.5–30
5-CH3-THF 0.17 0.53 36024.3 ± 682.2 0.998 1–30
5-CHO-THF 0.37 1.12 19204.0 ± 122.3 0.994 1–30
Table 2
Percentage recovery of spiked folate standards in tomato fruit
extract. 2.7 ng/mL of folate standards were spiked into extract.
Compound % recovery mean ± S.E.
FA 48.97 ± 8.48
THF 70.89 ± 7.10
5,10-CH+
THF 94.72 ± 8.29
5-CH3-THF 131.31 ± 8.78
5-CHO-THF 115.90 ± 1.59
MTX 95.27 ± 11.52
K. Tyagi et al. / Food Chemistry 179 (2015) 76–84 81
7. with earlier report (Iniesta, Perez-Conesa, Garcia-Alonso, Ros, &
Periago, 2009), THF was not detected in tomato fruit extract.
Tomato is consumed in both cooked and raw form by the people.
Taking the folate content of fully red ripe tomato fruit (18.06 lg/
100 g of FW) in account, its percentage contribution to the RDA
(400 lg/day) is 4.51. However, the folate content in a given food
matrix such as tomato fruit is not constant as several factor affects
its level such as nature of cultivar, developmental stage, and envi-
ronmental conditions during ripening (Iniesta et al., 2009;
Jägerstad et al., 2005).
The extent of basal polyglutamylation status of folate helps in
its retention in the cellular compartments as well as increases its
affinity toward the enzymes with which they interact (Akhtar
et al., 2010). The polyglutamylation of individual folate derivatives
was evaluated by comparing the monoglutamate amounts of each
folate derivative detected in tomato red ripe fruits with and with-
out the addition of rat plasma during folate extraction. 83% of 5-
CH3-THF was present in polyglutamylated form. Similarly, $90%
of total 5-CHO-THF was present in the polyglutamylated form
(Fig. 3C), whereas $50% 5,10-CH+
THF was present in polyglutamy-
lated form.
3.8. Application of the method
Fruits and vegetables are good sources of naturally occurring
folate, primarily 5-CH3-THF. To check the efficiency and applicabil-
ity of the above method for other plant materials, the folate levels
were determined in tomato juvenile leaf, spinach leaf, garden pea
seeds and capsicum fruit (Table 3). The major folate detected in
these varieties was 5-CH3-THF. Other forms observed were
5-CHO-THF and 5,10-CH+
THF with traces of THF and FA. 5-CHO-
THF was the second most predominant form detected in all of
them. Contrary to the report that recovery of THF was low at acidic
pH but increased as the pH of extraction buffer increased to 7.0
(Zhang et al., 2005), no THF was detected in tomato fruit tissue
using pH 7.0 extraction buffer. Compared to fruits, the juvenile
leaves of tomato showed very high level of folates, which may be
related to active cell division and expansion in these leaves.
Interestingly, folic acid or other oxidation products of natural folate
forms were not detected indicating that extraction procedure used
in this study led to minimal degradation or loss of folate.
The total folate content mean (±SE) in tomato juvenile leaf,
spinach leaf, garden pea seeds and capsicum fruit was 429.54
(61.49), 287.16 (55.14), 188.33 (14.06) and 39.44 (2.97) lg/100 g
FW respectively (Table 3). Considering that these values are higher
than the reported values in literature supports that our extraction
method leads to maximum recovery of folates from matrix. Total
folate content of spinach reported in earlier studies varied from
194–364 lg/100 g of FW assessed through microbiological assay
(Lin & Lin, 1999) to 48–177 lg/100 g of FW through HPLC
(Johansson, Jägerstad, & Frølich, 2007; Vahteristo, Lehikoinen,
Ollilainen, & Varo, 1997). Our results showed nearly 110 lg higher
value of folate than the earlier reported value for spinach. While
these differences in the folate levels could be attributed to cultivar,
Table 3
Folate content of tomato fruit, leaf and other vegetables (lg/100 g FW).
Sample THF (mean ± SE) 5-CH3-THF (mean ± SE) 5,10-CH+
THF (mean ± SE) 5-CHO-THF (mean ± SE) Total folate (mean ± SE)
Tomato fruit (mature green) – 14.97 ± 0.74 1.15 ± 0.08 5.81 ± 0.56 21.93 ± 1.17
Tomato fruit (red ripe) – 10.61 ± 1.42 1.55 ± 0.19 5.89 ± 0.59 18.05 ± 2.01
Tomato leaf 4.12 ± 0.10 226.36 ± 29.95 13.13 ± 2.92 185.93 ± 28.14 429.54 ± 61.49
Spinach 0.24 ± 0.09 159.17 ± 30.79 19.21 ± 4.05 108.54 ± 20.71 287.16 ± 55.14
Garden pea – 156.19 ± 7.62 4.28 ± 0.78 27.86 ± 2.67 188.33 ± 14.06
Green capsicum – 32.36 ± 2.76 2.06 ± 0.14 5.02 ± 0.55 39.44 ± 2.97
Fig. 3. Folate content in tomato fruits. (A) Total folate content in mature green (MG) and red ripe (RR) fruits (n P 3). (B) Relative distribution of different folate forms in
mature green and red ripe fruit (n P 3). (C) Polyglutamylation status of different folate derivatives in red ripe fruit.
82 K. Tyagi et al. / Food Chemistry 179 (2015) 76–84
8. variety, and geographical area, climatic and seasonal differences
among study groups (Shohag et al., 2011), it is also likely that sam-
ple preparation and extraction protocol reported in this study may
have led to more efficient extraction and minimal losses of folate
from spinach.
4. Conclusion
An accurate HPLC–MS/MS method has been successfully devel-
oped and validated for folate analysis in tomato fruits, which is
simple, fast and cost effective and can be applied to a large number
of samples for folate analysis. Our method has wide applicability as
the folate levels determined in different types of tissue used in the
study were either comparable or more than those reported in pre-
vious studies. We found that 5-CH3-THF is the main folate deriva-
tive in tomato. This method is fairly simple to execute as the
sample cleanup involves ultra-filtration instead of SPE or affinity
chromatography, without compromising on the amount or forms
of folates detected. The validation experiments showed linearity
of the method and total folate concentration in mature green and
red ripe tomato fruit was 21.93 and 18.06 lg/100 g of fresh weight
with acceptable precision and accuracy. We have used MTX as an
internal standard in screening of multiple samples for reproducible
quantitation. Most of the folates were stable for more than 24 h
when kept at 4 °C in autosampler. Total folate profile in tomato leaf
differed considerably from that of tomato fruit tissue indicating
that this extraction procedure can be used to analyze other food
matrices too.
Acknowledgments
This work was supported by the Department of Biotechnology
(Grant No. BT/PR11671/PBD/16/828/2008 to R.S. and Y.S.), the
Council of Scientific and Industrial Research (research fellowship
to KT), University Grants Commission (research fellowship to PU
and SS).
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.foodchem.2015.
01.110.
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