Effects of Smokeless Tobacco Samples from Tabuk Saudi Arabia on Nitric Oxide Production: A Potential Risk for Cancer and Cardiovascular Diseases

Background: Smokeless tobacco (SLT) is traditionally used in Middle East countries. The several toxic constituents with potential carcinogenicity make it a serious human health risk. Literature regarding their effects on cardiac and cancer disease is lacking in Saudi Arabia. Objective: This study was conducted to investigate the adverse effect of 11 different samples of widely used SLT varieties from the Tabuk region - Saudi Arabia, on nitric oxide (NO) level and their potential risk on cardiovascular health, etiology and/or progression of cancers. Methods: Samples were collected from Tabuk, KSA and analyzed by the GC-MS technique. Nitric oxide inhibition was performed using J774.2 macrophages by the Griess method. The retrieved crystallized structure of human inducible nitric oxide synthase (iNOS) from Brookhaven Protein Data Bank Repository PDB I.D: 3E7G with 2.20Å resolution was further prepared by structure using the MOE.2019 tool. The compounds abstracted from 11 different Shammah varieties were sketched by the MOE-Builder tool. Minimization for both receptor and compounds was performed via AMBER99 and MMFF99X force field implemented in MOE. Results: Nine samples (4 - 11) showed a potent suppressive effect on NO production with IC50 values ranging between (16.9-20.4 µg/mL), respectively. The samples (1 & 2) exhibited a moderate level of inhibition with IC50 ranging between 33.2 and 57.4 µg/mL, respectively. Interestingly, sample 4 consisting of compounds (13-15, 19-26, 28) that mostly belongs to the group fatty acid ester and phthalic acid ester showed the most potent suppressive effect. Molecular docking results revealed that the current local SLT constituents presented noticeable potency in different extract samples. Conclusion: Variable suppressive effects on NO were detected in the current SLT samples, where sample 4 was the most potent among all. The extract of the latter exhibited molecular interaction with the first shell amino acid residues of Inducible nitric oxide synthase (iNOS), which may anchor the plasticity and selectivity of the compounds present in it. The samples (4 -11) showed a potent inhibitory effect on the NO, where compound 26 (Phthalic acid ester) is common, and its adequate concentration may account for augmented biological activity. These results may effectively highlight their adverse effects on cardiovascular health and etiology and/or progression of cancer and may help in strengthening the social and governmental efforts in minimizing the use of these substances.

Degradation of dibutyl phthalate by Paenarthrobacter sp. Shss isolated from Saravan landfill, Hyrcanian Forests, Iran

Phthalic acid esters are predominantly used as plasticizers and are industrially produced on the million ton scale per year. They exhibit endocrine-disrupting, carcinogenic, teratogenic, and mutagenic effects on wildlife and humans. For this reason, biodegradation, the major process of phthalic acid ester elimination from the environment, is of global importance. Here, we studied bacterial phthalic acid ester degradation at Saravan landfill in Hyrcanian Forests, Iran, an active disposal site with 800 tons of solid waste input per day. A di-n-butyl phthalate degrading enrichment culture was established from which Paenarthrobacter sp. strain Shss was isolated. This strain efficiently degraded 1 g L–1 di-n-butyl phthalate within 15 h with a doubling time of 5 h. In addition, dimethyl phthalate, diethyl phthalate, mono butyl phthalate, and phthalic acid where degraded to CO2, whereas diethyl hexyl phthalate did not serve as a substrate. During the biodegradation of di-n-butyl phthalate, mono-n-butyl phthalate was identified in culture supernatants by ultra-performance liquid chromatography coupled to electrospray ionization quadrupole time-of-flight mass spectrometry. In vitro assays identified two cellular esterase activities that converted di-n-butyl phthalate to mono-n-butyl phthalate, and the latter to phthalic acid, respectively. Our findings identified Paenarthrobacter sp. Shss amongst the most efficient phthalic acid esters degrading bacteria known, that possibly plays an important role in di-n-butyl phthalate elimination at a highly phthalic acid esters contaminated landfill.

Biodegradation of Di (2-Ethylhexyl) Phthalate by a novel Enterobacter spp. Strain YC-IL1 Isolated from Polluted Soil, Mila, Algeria

Di-(2-ethylhexyl) phthalate (DEHP) is one of the phthalic acid ester representatives and is mainly used as a plasticizer to endow polyvinyl chloride plastics with desirable physical properties. It is synthesized in massive amounts worldwide. Many studies have proved the adverse effects of DEHP on human health and wildlife. DEHP is labeled as an endocrine disruptor which causes human reproductive problems. Enterobacter spp. YC-IL1, a novel isolated strain from contaminated soil, was identified by 16S rRNA gene analysis and electronic microscope. It is capable of efficiently degrading DEHP (100%) and a wide range of phthalic acid ester PAEs, particularly those containing side chains with branches, or ring structures such as dutylbenzyl phthalate and dicyclohexyl phthalate, which are hard to degrade, with, respectively, 81.15% and 50.69% degradation after 7 days incubation. YC-IL1 is an acido-tolerant strain which remained in pH values lower than pH 5.0 with the optimum pH 7.0 and temperature 30 °C. The DEHP metabolites were detected using HPLC-QQQ and then the degradation pathway was tentatively proposed. Strain YC-IL1 showed high DEHP degradation rate in artificially contaminated soil with 86% removed in 6 days. These results indicate the application potential of YC-IL1 in bioremediation of PAE-polluted sites, even the acidic ones.

The Impact of Storage on Phthalic Acid Esters Concentrations in Yogurts Packed in Plastic Cups

Phthalic acid esters are used as plastic softeners and also can be found in food packaging materials. European legislation defines specific migration limits of plastic additives for plastic materials that come into contact with food. This study monitors the phthalic acid ester concentrations in yogurts after manufacturing and then after a 3‑week storage. The studied yoghurts were natural yogurt with 1 % of chia flour, natural yogurt with 5 % of chia flour, natural yogurt with 1 % of bamboo fibre, natural yogurt with 5 % of bamboo fibre and natural yogurt. The analysed phthalic acid esters were dibutyl phthalate (DBP) and di‑(2‑ethylhexyl) phthalate (DEHP). The average phthalate concentrations in plastic cups were detected for DBP of 59.5 µg/g and for DEHP of 9.0 µg/g of the plastic material. Higher DBP concentrations than DEHP concentrations were also found in all studied yogurts. The average DBP concentrations in yogurts were detected from 1.8 µg/g up to 5.0 µg/g of the original matter and the average DEHP concentrations were determined from 0.5 µg/g up to 1.0 µg/g of the original matter. No statistically significant difference was found when comparing phthalic acid ester concentrations in yogurts immediately after production and after three weeks of storage. However, in our study in all cases of yogurts, the DBP concentrations were higher than the specific migration limit set by the legislation (0.3 mg/kg) and the DEHP concentrations were in all cases of yogurts lower than the specific migration limit set by the legislation (1.5 mg/kg).