Fumigation on bulk cargo ships: a chemical threat to seafarers.

Fumigation is a process that is carried out to prevent deterioration of goods by pests and spread of unwanted organisms for example during long-distance carriage by sea. Several intoxications due to use of pesticides on bulk cargo ships have been indicated, but for some of these incidents the documentation are questionable. The objective of the present study was therefore to examine the extent of the problem by collecting available information of incidents or intoxications due to use of pesticides on bulk cargo ships. Information sources such as PubMed, Google Scholar, Gard (marine insurance company), Marine Accident Investigation Branch, United Kingdom, and Professional Mariner (a magazine) were searched using similar search phrases. The results indicate that the present practice of fumigation with pesticides of cargo holds on bulk ships represents a serious health risk to both seafarers and port workers. A thorough search for information in both scientific and non-scientific sources revealed a number of intoxications including several fatalities. According to the available documentation, phosphine seems to be used more or less exclusively as fumigant on bulk cargo ships today. Phosphine has a high acute toxicity, and recent findings suggest long-term effects. Several of the reported incidents point to lack of knowledge and neglecting of recommended procedures as key elements in this respect. The problem is likely underestimated due to lack of available documentation of several incidents. Preventive actions should be implemented that focus on documentation of incidents, increase knowledge of pesticide health hazard and implementation of safety procedures that are mandatory to perform when fumigated cargo is to be handled on bulk ships.

Nitric oxide in exhaled gas and tetrahydrobiopterin in plasma after exposure to hyperoxia.

The fraction of nitric oxide in exhaled gas (FENO) is decreased after exposure to hyperoxia in vivo, although the mechanisms for this decrease is not clear. A key co-factor for nitric oxide synthase (NOS), tetrahydrobiopterin (BH4), has been shown to be oxidized in vitro when exposed to hyperoxia. We hypothesized that the decrease of FENO is due to decreased enzymatic generation of NO due to oxidation of BH4. The present study was performed to investigate the relationship between levels of FENO and plasma BH4 following hyperoxic exposure in humans. Two groups of healthy subjects were exposed to 100% oxygen for 90 minutes. FENO was measured before and 10 minutes (n = 13) or 60 minutes (n = 14) after the exposure. Blood samples were collected at the same time points for quantification of biopterin levels (BH4, BH2 and B) using LC-MS/MS. Each subject was his or her own control, breathing air for 90 minutes on a separate day. Hyperoxia resulted in a 28.6 % decrease in FENO 10 minutes after exposure (p < 0.001), confirming previous findings. Moreover, hyperoxia also caused a 14.2% decrease in plasma BH4 (p = 0.012). No significant differences were observed in the group measured 60 minutes after exposure. No significant correlation was found between the changes in FENO and BH4 after the hyperoxic exposure (r = 0.052, p = 0.795), this might be due to the recovery of BH4 being faster than the recovery of FENO.

Hyperoxia and lack of ascorbic acid deplete tetrahydrobiopterin without affecting NO generation in endothelial cells.

Nitric oxide (NO) may protect against gas bubble formation and risk of decompression sickness. We have previously shown that the crucial co-factor tetrahydrobiopterin (BH4) is oxidized in a dose-dependent manner when exposed to hyperoxia similar to diving conditions but with minor effects on the NO production by nitric oxide synthase. By manipulating the intracellular redox state, we further investigated the relationship between BH4 levels and production of NO in human endothelial cells (HUVECs). HUVECs were cultured with and without ascorbic acid (AA) and the glutathione (GSH) synthesis inhibitor buthionine sulfoximine, prior to hyperoxic exposure. The levels of biopterins and GSH were determined in cell lysates while the production of NO was determined in intact cells. Omitting AA resulted in a 91% decrease in BH4 levels (0.49 ± 0.08 to 0.04 ± 0.01 pmol/106 cells, p⟨0.001) at 20 kPa oxygen (O2), and 88% decrease (0.24 ± 0.03 to 0.03 ± 0.01 pmol/106 cells, p=0.01) after exposure to 60 kPa O2. The NO generation was decreased by 23% (74.5 ± 2.2 to 57.3 ± 5.6 pmol/min/mg protein, p⟨0.001) at 20 kPa O2, but no significant change was observed at 60 kPa O2. GSH depletion had no effects on the NO generation. No correlation was found between NO generation and the corresponding intracellular BH4 concentration (p=0.675, r=-0.055) or the BH4 to BH2 ratio (p=0.983, r=0.003), determined across 18 in vitro experiments. Decreased BH4 in HUVECs, due to hyperoxia or lack of ascorbic acid, does not imply corresponding decreases in NO generation.

Decrease of tetrahydrobiopterin and NO generation in endothelial cells exposed to simulated diving.

Purpose: Nitric oxide (NO) has been shown to protect against bubble formation and the risk of decompression sickness. We hypothesize that oxidation of tetrahydrobiopterin (BH4) leads to a decreased production of NO during simulated diving. Methods: Human umbilical vein endothelial cells (HUVEC) were exposed to hyperoxia or simulated diving for 24 hours. The levels of biopterins (BH4, BH2 and B) were determined by LC-MS/MS, and the production of NO by monitoring the conversion of L-arginine to L-citrulline. Results: Exposure to hyperoxia decreased BH4 in a dose-dependent manner; by 48 ± 15% following exposure to 40 kPa O2 (P⟨0.001 vs. control at 20 kPa O2), and 70 ± 16% following exposure to 60 kPa O2. Exposure to 40 kPa O2 decreased NO production by 25 ± 9%, but there was no further decrease when increasing oxygen exposure to 60 kPa (25 ± 10%). No additional effects of simulated diving were observed, indicating no additive or synergistic effects of hyperbaria and hyperoxia on the BH4 level or NO generation. Conclusion: NO generation in intact human endothelial cells was decreased by simulated diving, as well as by hyperoxic exposure, while BH4 levels seem to be affected only by hyperoxia. Hence, the results suggest that BH4 is not the sole determinant of NO generation in HUVEC.

Diving exposure and health effects in divers working in different areas of professional diving.

BACKGROUND: The aim of the present study was to compare diving exposure and health effects in different areas of professional diving. MATERIALS AND METHODS: The Norwegian Labour Inspection Authority's Diving Register contains data on all professional inshore divers who have held a diving certificate at any time since 1980. Of these divers, the "Norwegian diver 2011" questionnaire was completed by 2848 (48.7%). A total of 1167 male divers reported that they often worked in one area of diving only (rescue diving, diving instruction, fish farming, quay/construction work and offshore/oil related). In the analysis of these divers, rescue divers were used as referents as they reported the lowest number of dives. RESULTS: Age distribution, the proportion of retired divers and the mean number of dives completed varied between the different areas of professional diving. Compared to rescue divers, divers in fish farming, quay/ /construction work and offshore/oil related work more often experienced physically demanding diving. Divers in fish farming more often had no day off after 3 days of physically demanding work compared to rescue divers. All groups except offshore divers reported making further dives after one physically demanding dive on the same day. All groups reported more frequent decompression sickness than did the referents and divers in quay/construction and offshore/oil related diving reported more frequent episodes of unconsciousness during diving than did the referents. Divers in fish farming, in quay/construction work and oil/ /offshore related diving obtained a higher symptom score than the referents and the two latter groups also reported more frequent adverse health effects due to diving than the referents. Health related physical and mental component summary scores were lower in all other groups than in referents. CONCLUSIONS: Compared to the rescue divers, divers in quay/construction work and offshore/oil related divers reported more adverse health effects and obtained a higher symptom score.

Health risks in international container and bulk cargo transport due to volatile toxic compounds.

To ensure the preservation and quality of the goods, physical (i.e. radiation) or chemical pest control is needed. The dark side of such consents may bear health risks in international transport and production sharing. In fact, between 10% and 20% of all containers arriving European harbors were shown to contain volatile toxic substances above the exposure limit values. Possible exposure to these toxic chemicals may occur not only for the applicators but also the receiver by off gassing from products, packing materials or transport units like containers. A number of intoxications, some with lethal outcome, occur not only during the fumigation, but also during freight transport (on bulk carriers and other transport vessels), as well as in the logistic lines during loading and unloading. Risk occupations include dock-workers, seafarers, inspectors, as well as the usually uninformed workers of importing enterprises that unload the products. Bystanders as well as vulnerable consumers may also be at risk. Ongoing studies focus on the release of these toxic volatile substances from various goods. It was shown that the half-lives of the off-gassing process range between minutes and months, depending on the toxic substance, its chemical reactivity, concentration, the temperature, the contaminated matrix (goods and packing materials), and the packing density in the transport units. Regulations on declaration and handling dangerous goods are mostly not followed. It is obvious that this hazardous situation in freight transport urgently requires preventive steps. In order to improve awareness and relevant knowledge there is a need for more comprehensive information on chemical hazards and a broader implementation of the already existing regulations and guidelines, such as those from ILO, IMO, and national authorities. It is also necessary to have regular controls by the authorities on a worldwide scale, which should be followed by sanctions in case of disregarding regulations. Further, fumigated containers must have a warning sign corresponding to international recommendations and national regulations, and freight documents have to indicate any potential hazard during stripping the goods.

Mortality among retired offshore divers in Norway.

BACKGROUND: Health effects of diving have been observed in divers who have not experienced any diving related accidents. The aim of the study was to study total, and cause specific mortality, in Norwegian retired professional offshore divers. MATERIALS AND METHODS: We carried out a mortality follow-up from 1997 to 2013 in a group of men, born 1930-1973. The diving cohort consisted of 386 male professional divers diving in the North Sea in the pioneer period from 1965 to 1990, of which 25 were dead. 1,467,769 Norwegian males were used as referents. This population was linked to the Norwegian Cause of Death Registry. RESULTS: Twenty five (6.5%) professional offshore divers had died. No differences were observed between divers and referents for overall mortality or for non-violent deaths, adjusted for year of birth. CONCLUSIONS: In this cohort of retired North Sea divers, the mortality pattern did not differ from that of the expected mortality.

Hyperoxia but not ambient pressure decreases tetrahydrobiopterin level without affecting the enzymatic capability of nitric oxide synthase in human endothelial cells.

Nitric oxide (NO) seems to be related to bubble formation and endothelial dysfunction resulting in decompression sickness. Bubble formation can be affected by aerobic exercise or manipulating NO. A prior heat stress (HS) has been shown to confer protection against decompression sickness in rats. An important question was if the oxidative environment experienced during diving limits the availability of the nitric oxide synthase (NOS) cofactor tetrahydrobiopterin (BH4). Human endothelial cells were used to investigate how HS and simulated diving affected NO synthesis and defense systems such as heat shock protein 70 (HSP70) and glutathione (GSH). BH4 was measured using a novel LC-MS/MS method and NOS by monitoring the conversion of radiolabeled L-arginine to L-citrulline. Increased pO2 reduced BH4 levels in cells in a dose-dependent manner independently of high pressure. This effect may result in decreased generation of NO by NOS. The BH4 decrease seemed to be abolished when cells were exposed to HS prior to hyperoxia. NOS enzyme was unaffected by increased pO2 but substantially reduced after HS. The BH4 level seemed to a minor extent to be dependent upon GSH and probably to a higher degree dependent on other antioxidants such as ascorbic acid. A simulated dive at 60 kPa O2 had a potentiating effect on the heat-induced HSP70 expression, whereas GSH levels were unaffected by hyperoxic exposure. HS, hyperoxia, and dive affected several biochemical parameters that may play important roles in the mechanisms protecting against the adverse effects of saturation diving.

Simultaneous quantification of tetrahydrobiopterin, dihydrobiopterin, and biopterin by liquid chromatography coupled electrospray tandem mass spectrometry.

A simple and rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based method was developed for the quantification of tetrahydrobiopterin (BH4), dihydrobiopterin (BH2), and biopterin (B) in human umbilical vein endothelial cells (HUVECs). Freshly prepared cell samples were treated with a mixture consisting of 0.2M trichloroacetic acid (TCA) and a cocktail of various antioxidants in order to precipitate proteins and other cellular components and to stabilize red/ox conditions in the lysates. Chromatography of the cell lysates was performed on a Poroshell 120 SB-C18 column (2.7µm, 150×2.1mm) using a stepwise gradient elution made from two mobile phases. Quantification was performed on a triple quadrupole mass spectrometer employing electrospray ionization with the operating conditions as multiple reaction monitoring (MRM) at positive ion mode. Total chromatographic run time was 23min. The method was validated for analysis in HUVECs, and the limits of quantification were 1nM for BH4 and BH2 and 2.5nM for B. Standard curves were linear in the concentration ranges of 1 to 100nM for BH4 and BH2 and 2.5 to 100nM for B. The current study reports a novel method for the simultaneous and direct quantification of BH4, BH2, and B in a single injection.

Proposal on limits for chemical exposure in saturation divers' working atmosphere: the case of benzene.

Saturation diving is performed under extreme environmental conditions. The divers are confined to a limited space for several weeks under high environmental pressure and elevated oxygen partial pressure. At present, divers are protected against chemical exposure by standard exposure limits only adjusted for the increased exposure length, i.e. from 8 to 24 hours a day and from 5 to 7 days a week. The objective of the present study was to indicate a procedure for derivation of occupational exposure limits for saturation diving, termed hyperbaric exposure limits (HEL). Using benzene as an example, a procedure is described that includes identification of the latest key documents, extensive literature search with defined exclusion criteria for the literature retrieved. Hematotoxicity and leukemia were defined as the critical effects, and exposure limits based upon concentration and cumulative exposure data and corresponding risks of leukemia were calculated. Possible interactions of high pressure, elevated pO2, and continuous exposure have been assessed, and incorporated in a final suggestion of a HEL for benzene. The procedure should be applicable for other relevant chemicals in the divers' breathing atmosphere. It is emphasized that the lack of interactions from pressure and oxygen indicated for benzene may be completely different for other chemicals.

Differential effects on nitric oxide synthase, heat shock proteins and glutathione in human endothelial cells exposed to heat stress and simulated diving.

Decompression sickness (DCS) may result from damage to the endothelium caused by the gas bubbles formed during decompression and may be related to nitric oxide (NO) production by nitric oxide synthase (NOS). Heat stress prior to diving has been shown to protect animals from DCS, and by simulating this treatment in human endothelial cells (HUVEC) we have shown that a simulated dive performed subsequent to a heat stress potentiated the heat-induced expression of HSP70 and increased the level of the antioxidant glutathione (GSH). Since operational saturation diving is performed at an increased oxygen level, HUVEC have been exposed to heat stress and simulated diving at 40 kPa O(2), comparing the response on HSP70, HSP90 and GSH level to the effects previously observed at 20 kPa O(2). In addition, we wanted to investigate the effect on both endothelial NOS (eNOS) protein and enzymatic activity. The present results showed that a heat stress (45°C, 1 h) decreased the NOS activity and the protein markedly. Hyperoxia (40 kPa) alone or a dive either at 20 or 40 kPa O(2),had no effects on NOS activity or protein. At 40 kPa O(2) a simulated dive after heat stress potentiated the HS-induced HSP70 response, whereas the heat-induced HSP90 response decreased. GSH levels were found to be inversely related to NOS activity and protein expression, and might be explained by a possible post-translational regulation by glutathionylation of eNOS protein. The results add to the limited knowledge of these critical factors in cellular defence mechanisms that can prevent injury during decompression.

Low-dose exposure to alkylphenols adversely affects the sexual development of Atlantic cod (Gadus morhua): acceleration of the onset of puberty and delayed seasonal gonad development in mature female cod.

Produced water (PW), a by-product of the oil-production process, contains large amount of alkylphenols (APs) and other harmful oil compounds. In the last 20 years, there have been increasing concerns regarding the environmental impact of large increases in the amounts of PW released into the North Sea. We have previously shown that low levels of APs can induce disruption of the endocrine and reproductive systems of Atlantic cod (Gadus morhua). The aims of this follow-up study were to: (i) identify the lowest observable effect concentration of APs; (ii) study the effects of exposure to real PW, obtained from a North Sea oil-production platform; and (iii) study the biological mechanism of endocrine disruption in female cod. Fish were fed with feed paste containing several concentrations of four different APs (4-tert-butylphenol, 4-n-pentylphenol, 4-n-hexylphenol and 4-n-heptylphenol) or real PW for 20 weeks throughout the normal period of vitellogenesis in Atlantic cod from October to January. Male and female cod, exposed to AP and PW, were compared to unexposed fish and to fish fed paste containing 17ß-oestradiol (E(2)). Approximately 60% of the females and 96% of the males in the unexposed groups were mature at the end of the experiment. Our results show that exposure to APs and E(2) have different effects depending on the developmental stage of the fish. We observed that juvenile females are advanced into puberty and maturation, while gonad development was delayed in both maturing females and males. The AP-exposed groups contained increased numbers of mature females, and significant differences between the untreated group and the AP-treated groups were seen down to a dose of 4 µg AP/kg body weight. In the high-dose AP and the E(2) exposed groups, all females matured and no juveniles were seen. These results suggest that AP-exposure can affect the timing of the onset of puberty in fish even at extremely low concentrations. Importantly, similar effects were not seen in the fish that were exposed to real PW.

Simulated diving after heat stress potentiates the induction of heat shock protein 70 and elevates glutathione in human endothelial cells.

Heat stress prior to diving has been shown to confer protection against endothelial damage due to decompression sickness. Several lines of evidence indicate a relation between such protection and the heat shock protein (HSP)70 and HSP90 and the major cellular red-ox determinant, glutathione (GSH). The present study has used human endothelial cells as a model system to investigate how heat stress and simulated diving affect these central cellular defense molecules. The results demonstrated for the first time that a simulated dive at 2.6 MPa (26 bar) had a potentiating effect on the heat-induced expression of HSP70, increasing the HSP70 concentration on average 54 times above control level. In contrast, a simulated dive had no significant potentiating effect on the HSP90 level, which might be due to the higher baseline level of HSP90. Both 2 and 24-h dive had similar effects on the HSP70 and HSP90, suggesting that the observed effects were independent of duration of the dive. The rapid HSP response following a 2-h dive with a decompression time of 5 min might suggest that the effects were due to compression or pressure per se rather than decompression and may involve posttranslational processing of HSP. The exposure order seemed to be critical for the HSP70 response supporting the suggestion that the potentiating effect of dive was not due to de novo synthesis of HSP70. Neither heat shock nor a simulated dive had any significant effect on the intracellular GSH level while a heat shock and a subsequent dive increased the total GSH level approximately 62%. Neither of these conditions seemed to have any effect on the GSH red-ox status.

Glutathione in blood cells decreases without DNA breaks after a simulated saturation dive to 250 msw.

INTRODUCTION: Saturation diving involves exposure to high pressure and elevated oxygen level. The impact of cellular defense systems like glutathione in protecting cells against oxidative DNA damage seems unclear. The aim of the present study was, therefore, to investigate whether diving conditions would affect blood cell glutathione and thus alter the mononuclear cells' (MNC) susceptibility to oxidative DNA damage. METHODS: Eight subjects participated in a simulated saturation dive to 2.6 MPa (250 msw) lasting 19.3 d (0.8 d compression, 6.6 d bottom phase, 11.9 d decompression) breathing helium-oxygen with PO2 ranging from 35 to 70 kPa (3.5-7.0 msw). Blood samples collected before compression and after decompression were analyzed for glutathione content and single-stranded DNA breaks. RESULTS: The results demonstrate for the first time that a simulated saturation dive decreased glutathione content in peripheral blood cells (32% decrease in MNC), and that the decrease was most pronounced in the erythrocytes (45%). Remarkably, no single-stranded DNA breaks could be detected in the MNC despite the low glutathione level. DISCUSSION: The results suggest that glutathione is a useful indicator of oxidative stress and that a low glutathione level represents no significant harm to the blood cells in the absence of other toxic agents. The lack of DNA strand breaks suggests that protection against oxidative DNA damage was mainly provided by mechanisms other than the glutathione system. Although previous investigations point to hyperoxia as the most plausible explanation for the present observations, the effect of high pressure cannot be excluded.

Additive toxicity of limonene and 50% oxygen and the role of glutathione in detoxification in human lung cells.

Limonene has many commercial applications and has been introduced as an environmentally acceptable solvent replacing halogenated hydrocarbons. Occupational exposure to limonene presumably occurs simultaneously with other chemicals including oxidative agents and may exert a heavy strain on cellular detoxifying capacity resulting in synergistic effects. The present study used oxygen as an example of an ubiquitous oxidative and radical forming agent and investigated the combination effects with limonene on human lung cells. Mechanistic information was gained by comparing the toxicity of limonene with a major oxidation product, limonene 1,2-epoxide, and by the involvement of glutathione in cellular detoxification. At cell culture conditions most similar to the in vivo situation oxygen did not increase the toxicity of limonene beyond an additive effect. The results further indicated that limonene 1,2-epoxide was not the active compound in limonene toxicity. Experimental evidence suggests that detoxification of limonene in human lung cells primarily occurs by mechanisms not involving the glutathione system and point to possible long-term effects of limonene exposure. The present knowledge indicates clearly that the mechanism of action of limonene on biological systems and particularly in combination with oxidative compounds still remains to be elucidated. In light of the frequent exposure of humans to such combinations further investigations into this issue are highly recommended.