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Volume 6, Issue 1, February Issue - 2018, Pages:230-235

Authors: Noor Daifallah Al-Goufi, Dr. Hala Salim Sonbol*
Abstract: Capparis cartilaginea is a shrub plant which used in traditional medicine to cure various diseases. Phytochemical analysis of Capparis cartilaginea plants extracts revealed the presence of isothiocyanates and flavonoids.  A dose dependent study was designed to assess effects of Capparis cartilaginea fruit extract on serum parathyroid hormones (PTH) and 1α,25-Dihydroxyvitamin D3 (1α,25(OH)2D3) levels in adult male and female Wistar rats, which could describe a regulation effect on bone metabolism. Healthy adult Wistar rats (20 female and 20 male) were treated with 1000, 2000 and 3000 mg/kg body weight (BW) of fruit extract via gavage for 6 weeks. The rats were housed under standard laboratory conditions (22±1ºC and 60% humidity) for 2 weeks prior to the experiment. Blood samples were collected to determine the level of PTH and 1α,25(OH)2D3 in serum. Results of study revealed that serum PTH levels were increased in male and female rats treated with 1000, 2000 and 3000 mg/kg body Capparis cartilaginea fruit extract.  The analysis of the hormones suggested a dose-dependent response in male serum PTH levels and it was significantly higher than the female PTH levels. On the other hand, all the three doses of Capparis cartilaginea fruit extract decreased 1α,25(OH)2D3 levels in both genders. The results of study indicated that Capparis cartilaginea fruit extracts have a potential to change the PTH and 1α,25(OH)2D3 hormonal levels.
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Full Text: 1 Introduction Traditional medicine covers a wide variety of therapies and practices, which vary from country to country and region to region (World Health Organization, 2013). The flora of Saudi Arabia is considered as the richest biodiversity area in the Arabian Peninsula that comprise important genetic resources of crops, medicinal plants and xerophytic vegetation which make up the prominent features of plant life in the kingdom (Zahran, 1982; Al-Yahya, 1984). According to Al-Yahya (1984), the Arabian Peninsula is the birthplace of herbal drugs and folk medicine. In addition to its large number of endemic species, the components of the flora are the admixture of the elements of Asia, Africa and Mediterranean region. Saudi Arabia is gifted with a wide range of flora, consisting of a large number of medicinal herbs, shrubs and trees. Saudi Arabian flora is expected to have more than 1200 medicinal species out of 2250 species in the flora (Mossa et al., 1987). Three hundred species have medicinal use (Rahman et al., 2004). Studies stated that about 24% of plants are medicinal in 15 families of which 30.1% are rare or threatened (Rahman et al., 2004; Yusuf et al., 2014). The total recorded vascular genera for the flora of Saudi Arabia stands at 855 and the number of species at 2,290, rising by ~ 2 % species throughout the past decade (Basahi et al., 2015). Medicinal plants represent important health and economic components of biodiversity. In each country, it is essential to conduct an inventory of medicinal components of the flora, for conservation and sustainable use (Seighali & Zaker, 2010). The uses of plants in Saudi Arabia for the cure of many illnesses are ancient and still available among the tribal and local people and traditional healers (Hakim) (Rahman et al., 2004). Among natural health products, Capparis cartilaginea                    (family Capparidaceae), has been found in the Saudi Arabian flora (Rahman et al., 2004). It has been used as important medicinal plants against various human diseases such as rheumatism, gout, paralysis, treating enlarged spleen and tuberculosis (Said, 1969; Nadkarni 1976; Al-Shayeb, 2012). It has also been reported that the crude extract of whole plant of Capparis cartilaginea produces a dose-dependent decrease in blood pressure and slight bradycardic rhythm in anaesthetised rats (Gilani & Aftab, 1994).  The presence of various phytochemicals has been reported from the crude extract of Capparis cartilaginea, among these isothiocyanates is most commonly reported one (Hamed et al., 2007; Al-Shayeb, 2012).Therapeutic use of isothiocyanates is also well reported and it can be used to treat arthritis and reduce the inflammatory status of synovium without disrupting the cellular                   homeostasis (Balar & Nakum, 2010). Rutin and quercetin are flavonoids that have been found in Capparis cartilaginea extract           (Ahmed et al.,1972; Sharaf et al., 1997). Beneficial health effects of these flavonoids are well reported and are known to have anticarcinogenic (Webster et al., 1996),                   anti-inflammatory, analgesic (Pietta & Gardana, 2003) and              anti-mutagenic properties (Brindzova et al., 2009), in addition to this, it has partial protective effect against the development of diabetes (Srinivasan et al., 2005). These chemicals were also used to treat skin inflammation, bruises, swellings, rheumatism, joint inflammation, knee problems, tendinitis, sprains, muscular contractions, paralysis, headaches, and earaches (Rivera et al., 2003). The role of phyto-flavonoids in regulation of various hormones such as estrogens, androgens and thyroid is well reported (Narayana et al., 2001). This regulation in humans is performed by binding to 17 beta-hydroxysteroid dehydrogenases, which in turn controls estrogen and androgen levels. Furthermore, it binds to beta-hydroxysteroid dehydrogenase, as a step towards regulating progestin and androgen levels (Noro et al.,1983). Furthermore, Quercetin and rutin influence the transport, metabolism and function of thyroid hormones (Tripathi & Rastogi, 1981).The current study has been undertaken to evaluate the impacts of different doses of ethanolic fruit extract of Capparis cartilaginea on the levels of PTH and 1α,25(OH)2D3 in adult Wistar male and female rats. PTH and the active form of vitamin D; 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], (also called calcitriol) control the mineral fluxes through the intestine, bone, kidney and blood (Favus et al., 2006). PTH promotes the reabsorption of Ca2+ from the bone into the circulation. In the kidney, it stimulates Ca2+ reabsorption and inorganic phosphate excretion in the urine.  PTH induces the hydroxylation of                  25-hydroxyvitamin D at the 1-position, forming the active form of vitamin D (calcitriol).  In intestine, absorption of dietary Ca2+ increases by increasing the level of Vitamin D. It also enables the renal reabsorption of filtered Ca2+. In the bones, vitamin D releases Ca2+ into the circulation by increasing bone reabsorption. Consequently, bone reabsorption is inhibited and the action of calcitonin, which amplifies the renal Ca2+ excretion. The communications between PTH, vitamin D and calcitonin leads to the maintenance of normal concentration of Ca2+ in blood plasma (Molina, 2013). Present study has been carried out to access the effect of Capparis cartilaginea fruit extract on the levels of these two hormones. 2 Materials and Methods 2.1 Animals In the present study, 20 males and 20 females (3-month-old; ~300g) adult Wister rats were obtained from the Animal House Unit at King Fahad Medical Research Centre (KFMRC), Jeddah, Saudi Arabia. The rats were stratified into four groups of each gender based on the dose of Capparis cartilaginea fruit extract (control, 1000, 2000 and 3000 mg/kg BW). The rats were accommodated under standard laboratory conditions (22±1ºC and 60% humidity) for 2 weeks prior to the experiment. They were under 12 h dark-light cycle (lights on at 0700 h), given a slandered pelleted diet (Grain Soils and Flour Mills Organization Jeddah, Saudi Arabia), with free access to water. Animals received care according to institutional guidelines for the care and the use of laboratory animals in KFMRC. The Research Ethics Committee, Unit of Biomedical Ethics, KAU, and Jeddah, Saudi Arabia have approved the experimental protocol. 2.2 Plant Material The fresh fruits of C. cartelaginea were collected from Umluj Mountains in Tabuk province, Northwest Saudi Arabia. All the collected fruit were freeze-driedat -64°C under 5m Torr pressure and grounded by Waring blender (USA). The freeze-dried fruit (50g) was used for extraction purpose, and extraction was carried out by 70% ethanol for 6-8 hours at 70°C using Soxhlet apparatus (Sigma, USA). After extraction, the mixture was evaporated by a rotary evaporator (Hahnapor, USA) at 60°C, concentrated under reduced pressure (100 torr), and dried by the freeze dryer. The dried extract was stored at -20°C until it is used. 2.3 Acute Oral Toxicity Test Acute oral toxicity test of C. cartelaginea fruit extract was performed (Organisation for Economic Co-operation and Development - OECD – 420, 2008), to select a proper dose for oral gavage. Groups of animals were dosed using the fixed doses of 5, 50, 300, 2000 and 5000 mg/kg body weight (BW). 2.4 Experimental Design After 2 weeks of acclimatisation, the rats were divided into four equal groups (n=5). Group one (control) was administrated with 2 ml of distilled water via oral gavage once daily. Group two was administrated with 1000mg/kg BW of C. cartelaginea fruit extract. Group three was administrated with 2000mg/kg BW of C. cartelaginea fruit extract. Group four was administrated with 3000mg/kg BW of C. cartelaginea fruit extract. The extract was administrated daily to the rats using oral gavage from Sunday to Thursday for 6 weeks. After the treatment period, blood was collected via the intraorbital sinus (Parasuraman et al., 2010) of the rats, using a capillary tube (75mm, Hirschmanlaborgerate, Germany) under ether anaesthesia. The blood was withdrawn into a plain tube for serum preparation. A collected blood sample was centrifuged at 3000 rpm for                    15 min. The serum was then stored in adeep freezer                                at -80°C until further use. 2.5 Measurement of 1α,25(OH)2D3 The 1α,25(OH)2D3 was measured from the rats’ serum by competitive inhibition enzyme immunoassay technique using the commercial 1α,25(OH)2D3 ELISA kit (CUSABIO Biotech CO. Ltd, China). The analysis was carried out according to the manufacturer’s instructions ( 2.6 Measurement of PTH The levels of PTH was measured from the rats’ serum, employing the quantitative sandwich enzyme immunoassay technique by using commercially available PTH ELISA kit (CUSABIO Biotech CO. Ltd, China). The analysis was carried out according to the manufacturer’s instructions ( 2.7 Statistical Analysis The data were analysed using the Statistical Package for the Social Sciences program version 21 (SPSS 21). Weight difference and biochemical parameters were analysed using one-way ANOVA. Post hoc testing was performed for inter-group comparisons. Results were expressed as themean ± standard deviation (SD), and the level of significance was set at P<0.05. 3 Results The doses used in the acute oral toxicity test performed on the rats did not show any visible sign of toxicity. 3.1 Biochemical Parameters The effects of Capparis cartilaginea fruit extract on the hormones levels in the serum of the male and female rats are summarised in table 1. Briefly, there was a dose dependent increase in serum PTH levels in male groups treated with Capparis cartilaginea fruit extract when compared to control group. Capparis cartilaginea fruit extract at a dose of 1000, 2000 and 3000 mg/kg raised the PTH levels by 14.5±5.4 pg/ml, 19±7 and 24.5±7.8 pg/ml respectively. However, the differences between various treatment are not statistically significant (p>0.05). The serum PTH levels increased in female groups treated with Capparis cartilaginea fruit extract as compared to the control group. A dose of 1000, 2000 and 3000 mg/kg of Capparis cartilaginea fruit extract raised the PTH levels to 12±4, 10±5 and 17.7±9.5 pg/ml respectively. This is also not showing statistically significant differences (p>0.05).   Table 1 Effect of oral administration of Capparis cartilaginea fruit extract on serum PTH and 1α,25(OH)2D3 in Wistar rats  
Groups   Female Male PTH (pg/ml) P-value 1α,25(OH)2D3 (pg/ml) P-value PTH (pg/ml) P-value 1α,25(OH)2D3 (pg/ml) P-value Group one (Control) 8±2.4 Ref 34.4±3.6 Ref 11.6±2.37 Ref 23.6±4.5 Ref Group two (1000 mg/kg) 12.0±4.1 0.67 33.5±3.60 0.98 14.5±5.4 0.99 23.4±1.8 0.99 Group three (2000 mg/kg) 10.30±5.1 0.92 28.3±4.3 0.1 19.0±7.0 0.23 22.3±1.9 0.89 Group four (3000 mg/kg) 17.70±9.5 0.09 27.9±2.80 0.07 24.5±7.8 0.63 18.7±2.1 0.06
Data are expressed as mean±SD, SD: standard deviation, PTH: parathyroid hormone, 1α,25(OH)2D3: 1 α,25-Dihydroxyvitamin D3
A dose of 1000 mg/kg Capparis cartilaginea fruit extract did not change the mean serum 1α,25(OH)2D3 levels (23.4 vs 23.6 pg/ml respectively) of group two male rats when compared to control, as shown in table 1. However, as the dose of Capparis cartilaginea fruit extract increase in male group three and four (2000 and 3000 respectively), the mean serum 1α,25(OH)2D3 levels decrease compared to control group (22.3, 18.7 vs 23.6 pg/ml respectively).   The reduction in the mean was more apparent in the female groups treated with Capparis cartilaginea fruit extract as compared to control group. A dose of 1000, 2000 and 3000 mg/kg Capparis cartilaginea fruit extract reduced the mean serum 1&

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