Degradation of estrogenic hormones in a silt loam soil

Estrogenic hormones are endocrine-disrupting compounds, which disrupt the endocrine system function of animals and humans by mimicking and/or antagonizing endogenous hormones. With the application of sludge biosolid and animal manure as alternative fertilizers in agricultural lands, estrogens enter the soil and become an environmental concern. The degradation kinetics of 17beta-estradiol, an estrogenic hormone of major concern, in a silt loam soil were investigated in this study. It was found that 17beta-estradiol degraded rapidly in nonsterilized soil with a half-life of 0.17 day. The degradation rate constant was proportional to the percentage of nonsterilized soil, indicating that microorganisms are directly responsible for the rapid degradation of 17beta-estradiol in soil. The half-life of 17beta-estradiol in 20% nonsterilized soil was slightly shortened from 1.3 to 0.69 day with the increase of soil moisture from 10 to 20% and was greatly decreased from 4.9 to 0.92 day with the increase of temperature from 15 to 25 degrees C. The coexistence of 40 micromol kg (-1) sulfadimethoxine, a veterinary antibiotic, decreased the degradation rate constant of 17beta-estradiol from 0.750 +/- 0.038 to 0.492 +/- 0.016 day (-1). The degradation kinetics of another three estrogenic hormones, including 17alpha-estradiol, estrone, and estriol, were also investigated and compared. Estrone was identified as a degradation product of 17beta-estradiol and the most persistent hormone among the four investigated estrogens. Estriol was observed in the degradation of estrone and 17alpha-estradiol.     

Naturally occurring estrogens in processed milk and in raw milk (from gestated cows)

The occurrence of the steroid hormones estrone (E1), 17alpha-estradiol (alphaE2), 17beta-estradiol (betaE2), and estriol (E3) in processed bovine milk with different fat contents and in raw milk from (non)gestated cows was investigated. Following liquid extraction, optional enzymatical deconjugation, C18 solid-phase extraction, and derivatization, estrogens were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Free and deconjugated E1 (6.2-1266 ng/L) was the major estrogen followed by alphaE2 (7.2-322 ng/L) and betaE2 (5.6-51 ng/L), whereas E3 was detected regularly at the detection limit of 10 ng/L. The lowest and highest concentrations were determined in raw milk from nonpregnant and from cows in the third trimester of gestation, respectively. The estrogen concentration in processed milk coincides with that of raw milk between first and second trimesters, reflecting the contribution of lactating pregnant cows to the final consumable product. The daily intake of total investigated estrogens through milk is 372 ng, which is dramatically more than currently recognized.     

SPE-HPLC/FLD法同时测定水中4种雌激素

研究建立了固相萃取（SPE）-高效液相色谱仪（HPLC）-荧光检测器（FLD）测定水体中4种雌激素（雌三醇、17β-雌二醇、炔雌醇和双酚A）的分析方法。水样过C18固相萃取柱净化浓缩,用5.00mL超纯水淋洗,15.00mL甲醇洗脱,洗脱液经氮气吹干后用50%甲醇溶解经HPLC-FLD测定;4种雌激素以甲醇/乙腈/水为流动相（体积比为25：30：45）,经InertsilODS-SP-C1（8150mm×4.6mm,5μm）反相色谱柱分离,激发和发射波长分别为280nm和310nm,流速1.0mL.min-1,柱温40℃,进样量20μL,以保留时间定性、外标法定量。该方法的线性范围为5.00～1000.00μg.L-1,且相关性良好（R〉0.9999）,4种雌激素的仪器检出限为0.107～0.271μg.L-1,方法检出限为0.214～0.540ng.L-1。在自来水中添加不同浓度的雌激素混合标准溶液,测得溶液中4种物质的加标回收率除炔雌醇为55.71%～66.78%外,其余雌激素的加标回收率均大于85%,相对标准偏差RSD（n=5）均小于4%。该方法灵敏度高、检出限低、重复性和精密性良好,能有效去除基质干扰,可用于水体中痕量雌激素的分析测定。     

Simultaneous determination of 20 pharmacologically active substances in cow's milk, goat's milk, and human breast milk by gas chromatography-mass spectrometry

This paper reports a systematic approach to the development of a method that combines continuous solid-phase extraction and gas chromatography-mass spectrometry for the simultaneous determination of 20 pharmacologically active substances including antibacterials (chloramphenicol, florfenicol, pyrimethamine, thiamphenicol), nonsteroideal anti-inflammatories (diclofenac, flunixin, ibuprofen, ketoprofen, naproxen, mefenamic acid, niflumic acid, phenylbutazone), antiseptic (triclosan), antiepileptic (carbamazepine), lipid regulator (clofibric acid), β-blockers (metoprolol, propranolol), and hormones (17α-ethinylestradiol, estrone, 17β-estradiol) in milk samples. The sample preparation procedure involves deproteination of the milk, followed by sample enrichment and cleanup by continuous solid-phase extraction. The proposed method provides a linear response over the range of 0.6-5000 ng/kg and features limits of detection from 0.2 to 1.2 ng/kg depending on the particular analyte. The method was successfully applied to the determination of pharmacologically active substance residues in food samples including whole, raw, half-skim, skim, and powdered milk from different sources (cow, goat, and human breast).     

Microbially Mediated Degradation of Common Pharmaceuticals and Personal Care Products in Soil Under Aerobic and Reduced Oxygen Conditions

Biological degradation rates of estrogen compounds and common pharmaceutical and personal care products (PPCPs) were examined in soils with a long history of exposure to these compounds through wastewater effluent and in soil not previously exposed. Biological degradation rates over 14 days were compared under aerobic and anaerobic conditions. Estrogen compounds including estrone, 17??-estradiol, estriol, and 17??-ethinylestradiol exhibited rapid degradation by soil microorganisms in both aerobic and anaerobic conditions. Rapid degradation rates for estrone, estriol, and 17??-ethinylestradiol occurred in pre-exposed soil under aerobic conditions; half-lives calculated under these conditions were 0.6, 0.7, and 0.8 day, respectively. Unexposed soil showed similar or slightly longer half-lives than pre-exposed soil under aerobic conditions. The exception was 17??-estradiol; in all treatments, degradation in unexposed soil resulted in a shorter half-life (2.1 versus 2.3 days). Anaerobic soils exhibited high biological degradation of estrogens as well. Half-lives of all estrogens ranged from 0.7 to 6.3 days in anaerobic soils. Triclosan degraded faster under aerobic conditions with half-lives of 5.9 and 8.9 days in exposed and unexposed soil. Under anaerobic conditions, triclosan half-lives were 15.3 days in unexposed and 28.8 days in exposed soil. Ibuprofen showed the least propensity toward biological degradation than other chemicals tested. Biological degradation of ibuprofen was only observed in unexposed soil; a half-life of 41.2 days was determined under anaerobic conditions and 121.9 days under aerobic conditions. Interestingly, unexposed soil exhibited a greater ability under anaerobic conditions to biologically degrade tested compounds than previously exposed soil.     

Adsorption of Natural Estrogens and Their Conjugates by Activated Sludge

Adsorption to biomass is a key mechanism which results in the elimination of natural estrogens and their conjugates from sewage. Freundlich model showed that the adsorption capacities of estrone and 17β-estradiol to activated sludge were the highest at neutral pH. The lower capacities at pH 2 and 11.5 could be due to the competition of sludge adsorption sites by cations or electrostatic repulsion from particles of similar charges. The lowest adsorption capacity at pH 11.5 was attributable to electrostatic repulsion, and the highest capacity at pH 2 might be due to the increased sulfate adsorbability. For estrogen conjugates such as estrone-3-sulfate and 17β-estradiol-3-sulfate, adsorption performances were similar at pH 5, 7, and 9. It was observed that mean values of log K _D were 2.78, 2.61, 1.67, and 1.94 l kg TSS^?1; log K _OM were 2.96, 2.79, 1.77, and 2.04 l kg VSS^?1 and those of log K _OC were 3.31, 3.12, 2.21, and 2.46 l kg OC^?1 for estrone, 17β-estradiol, estrone-3-sulfate, and 17β-estradiol-3-sulfate, respectively.     

Occurrence of PPCPs at a Wastewater Treatment Plant and in Soil and Groundwater at a Land Application Site

Pharmaceuticals and personal care products (PPCPs) can reach soil and aquatic environments through land application of wastewater effluent and agricultural runoff. The objective of this research was to assess the fate of PPCPs at field scale. PPCPs were measured systematically in a wastewater treatment plant (WWTP), and in soil and groundwater receiving treated effluent from the WWTP. A land application site in West Texas was used as the study site; it has received treated wastewater effluent from the WWTP for more than 70 years in order to remove additional nutrients and irrigate non-edible crops. Target compounds (estrone, 17??-estradiol, estriol, 17??-ethynylestradiol, triclosan, caffeine, ibuprofen, and ciprofloxacin) in wastewater, sewage sludge, soil, and groundwater were determined using HPLC/UV with qualitative confirmatory analyses using GC/MS. Samples were collected quarterly over 12 months for wastewater and sludge samples and over 9 months for soil and groundwater samples. Results indicated that concentrations of PPCPs in wastewater influent, effluent, sludge solid phase, and sludge liquid phase were in the range of non-detected (ND)-183 ??g/L, ND-83 ??g/L, ND-19 ??g/g, and ND-50 ??g/L, respectively. Concentrations in soil and groundwater samples were in the range of ND-319 ng/g and ND-1,745 ??g/L, respectively. GC/MS confirmation data were consistent with the results of HPLC/UV analyses. Overall, data indicate that PPCPs in the wastewater effluent from the WWTP transport both vertically and horizontally in the soil, and eventually reach groundwater following land application of the effluent.     

Occurrence of Sexual Hormones in Sediments of Mangrove in Brazil

The presence of sexual hormones (female estrogens) was assessed in sediments of a mangrove located in the urban region of southern Brazil. The estrogens are involved in human sexual reproduction. They act as the chemical messengers, and they are classified as natural and synthetic. The estrogens inputs in the environment are from treated and untreated sewage. The presence of estrogens in sewage is excretion from the female due to natural production and use of contraceptives (synthetic estrogens). With the indiscriminate release of sewage into the environment, estrogens can be found in rivers, lakes, and even in oceans. In this work, the presence of estrone (E1), 17-??-estradiol (E2), and 17-??-ethynilestradiol (EE2) in eight sedimentary stations in Itacorubi mangrove located on Santa Catarina Island, south Brazil, was investigated. Historically, the Itacorubi mangrove has been impacted by anthropogenic activities because the mangrove is inserted in the urban area of the Florianopolis. The estrogen EE2, used as contraceptive, had the highest concentration in mangrove sediment, 129.75?±?3.89 ng/g. E2 was also found, with its concentration ranging from 0.90?±?0.03 to 39.77?±?1.19 ng/g. Following the mechanism, under aerobic or anaerobic conditions, E2 will first be oxidized to E1, which is further oxidized to unknown metabolites and finally to CO₂ and water (mineralized). EE2 is oxidized to unknown metabolites and also finally mineralized. Theoretically, under anaerobic conditions, EE2 can be reduced to E1 even in environments such as mangrove which is essentially anaerobic.     

Degradation by Solar Radiation of Estrogenic Hormones Monitored by UV?CVisible Spectroscopy and Capillary Electrophoresis

This work aims to study the degradation of estrone, 17??-estradiol, 17??-ethinylestradiol, and estriol under direct solar radiation, with an average irradiance value of 5.2?kWh/m². Degradation at different temperatures (4??C, 20??C, and 30??C) was also tested in darkness. Individual solutions of the four estrogens were prepared and subjected to the conditions referred to above. The degradation for each compound was followed, after 7, 14, 21, 28, 35, 63, 91, and 126?days measuring the absorbance at the wavelength of 281?nm. The degradation of the four mixed estrogens was determined using capillary electrophoresis (CE), with a diode array detector and cholic acid, sodium salt plus sodium borate as a background buffer. The results showed no significant degradation rates on samples subjected to different temperatures. However, the results from CE analysis showed that, under direct solar radiation, after 126?days, the degradation rate varied between 75% and 100%. Also, the UV?CVis showed significant changes in the shape of UV?CVis spectra under direct solar radiation.     

Degradation of 17β-Estradiol and its Metabolites by Sewage Bacteria

Natural estrogens (e.g., 17β-estradiol or 1,3,5[10]-estratriene-3,17β-diol) have been suggested as one of the major groups of substances that cause endocrine disruption in wildlife. There is little information in the open literature on the fate of natural estrogens in the environment, a fact thathinders the assessment of their ultimate impact on the ecosystem. Aerobic and anaerobic batch experiments involving a 17β-estradiol-degrading culture and a supernatant of activated sludge from a local sewage treatment plant (Burlington, Ontario) were undertaken to assess the persistenceof 17β-estradiol (E₂) and its 5 metabolites. The batch experiments showed that E₂ and the metabolites werenot persistent and could be rapidly degraded by sewage bacteria.Biodegradation of E₂ by sewage bacteria appeared to initiate at the D ring of E₂, leading to the formation of the major metabolite estrone (E₁). No other major degradation products were noted. However, during the very earlystages of E₂ degradation by sewage bacteria, a previouslyunreported metabolite, X1 (5-hydroxy-15-methyl-13-oxatetracyclo[8.7.0.0 <2,7> .0. <11,15>]-heptadeca-2(7),3,5-trien-14-one), was observed. X1 appeared to be a labilemetabolite with a lactone structure, but its significance in thebiodegradation of E₂ remained to be elucidated. With theobservation of the new metabolite X1, a metabolic pathway of E₂ by sewage bacteria was proposed. Conditions (e.g., aerobic and anaerobic environment) governing the persistence of E₂ in sewage were also investigated. Results in this study suggest that the risk of extensive accumulation of natural estrogens normally found in sewage effluents in theenvironment is small, due to their ready biodegradation.     

Simultaneous determination and differentiation of glycidyl esters and 3-monochloropropane-1,2-diol (MCPD) esters in different foodstuffs by GC-MS

The aim of this study was the development of a method for the simultaneous determination and differentiation of fatty acid esters of 3-monochloropropane-1,2-diol (3-MCPD esters) and glycidol (glycidyl esters) in different foodstuffs. The esters were isolated from fat-rich food samples using a single extraction step and separated from interfering substances. For differentiation of 3-MCPD esters and glycidyl esters the glycidol moiety was converted into 3-methoxypropane-1,2-diol (3-MPD) by acidic alcoholysis. Subsequent determination was achieved by isotope dilution GC-MS after transesterification using an isotope-labeled 3-MCPD ester as internal standard. During optimization of the procedure, critical parameters affecting simultaneous determination and differentiation of these analytes were investigated. Rapid ester cleavage and derivatization at ambient temperature proved to be essential for the simultaneous determination of these analytes. The method was validated for various fat-rich foodstuffs such as bakery products, sweets, gravy, and soup powders as well as edible fats and oils. LODs of 8 and 15 μg/kg (fat-rich foodstuffs) as well as 50 and 65 μg/kg (edible oils and fats) were obtained for 3-MCPD esters and glycidyl esters, respectively. Recoveries for 3-MCPD esters and glycidyl esters ranged within 98 ± 4 and 88 ± 2% in all tested foodstuffs (0.05-2.5 mg/kg) and within 99 ± 16 and 93 ± 13% for edible oils and fats (0.15-3 mg/kg) over a wide concentration range. These results proved an accurate and differentiated determination of 3-MCPD esters and glycidyl esters with successful application to the fast screening of samples, avoiding tedious and laborious sample preparation.     

Simultaneous determination of chloramphenicol and aflatoxin M1 residues in milk by triple quadrupole liquid chromatography-tandem mass spectrometry

A reliable, rapid, and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous determination of chloramphenicol and aflatoxin M(1) in milk has been developed. This method includes simple extraction of sample with acetonitrile, separation on a MGIII-C(18) column using 5 mM ammonium acetate aqueous solution/methanol (60:40, v/v) as mobile phase, and MS/MS detection using multiple reaction monitoring mode. The method was validated according to Commission Decision 2002/657/EC. The limits of detection (LODs) were 0.05 μg/kg for chloramphenicol and 0.005 μg/kg for aflatoxin M(1.) The limits of quantification (LOQs) were 0.2 μg/kg for chloramphenicol and 0.02 μg/kg for aflatoxin M(1). The recovery values ranged from 88.8% to 100.6%, with relative standard deviation lower than 15% in all cases, when samples were fortified at three different concentrations. The decision limits (CCα) and detection capability (CCβ) of the method were also reported. This method has been successfully applied for simultaneous analysis of chloramphenicol and aflatoxin M(1) residues in milk from local supermarkets in China.     

Biodegradation of estrone and 17 β-estradiol in grassland soils amended with animal wastes

The release of endocrine disrupting chemicals into the environment is of increasing concern due to the formation of an intersex state in freshwater organisms and potential risks to human health. The aim of this study was to investigate the persistence of the naturally occurring hormones, estrone and 17 β-estradiol in three agricultural grassland soils in the presence and absence of cattle and sheep wastes (urine and manure). Biodegradation was investigated using ¹⁴C-labelled hormones which were applied to soil in three different solvents (water, artificial urine and natural sheep urine). When applied directly to soil the two hormones degraded at a similar rate, however, the speed of mineralization was soil type and solvent dependant. The half-life (t_1/2) of the hormones in soils ranged from 5 to 25 d. The hormones were also applied to the soils in sheep and cattle manure of different ages (7 d to 2 yr). Generally, the rate of degradation in the animal manure amended soils was more rapid than in the unamended soils (t_1/2=1-9 d), with mineralization being largely independent of manure age and type. We conclude that in comparison to many xenobiotics, estrogens are not persistent in agricultural soils. However, our calculations suggest that if they are lost to freshwater via runoff or leaching then they may have an appreciable effect on freshwater organisms. Assuming normal landspreading rates our results suggest that the risk of estrogen contamination of freshwater associated with manure spreading is very low.     

Effect of cooking on concentrations of β-estradiol and metabolites in model matrices and beef

Because beef food products are generally cooked prior to consumption, the behavior of chemicals in these cooked foods is important in estimating human exposure. The heat stability of the natural estrogen β-estradiol (β-E2) and its metabolites α-estradiol (α-E2), estrone (E1), and several catechol estrogens was examined in heated vegetable oil and aqueous solutions. The chemicals were also incorporated into regular and extra lean ground beef and subjected to cooking. E1 and E2 were stable in aqueous solutions at 100°C, whereas the catechol estrogens exhibited first-order decay curves with half-lives of 2-10 min. Their stability improved to the same level as the other test chemicals when an antioxidant was added to the solution, suggesting that their disappearance was due to oxidation rather than thermal degradation. E1 and E2 were also stable in heated vegetable oil (160-180°C), whereas catechol estrogen decreased 30-50% over the 2 h duration of the experiments. Chemical losses from cooked beef appear to be related to the fat content of the beef, with greater losses occurring in regular ground beef (25-30%), compared to extra lean ground beef (5-20%). This study shows that cooking reduces but does not eliminate the potential for dietary exposure to growth promoters in ground beef.     

Determination of furaltadone and nifursol residues in poultry eggs by liquid chromatography-electrospray ionization tandem mass spectrometry

The use of nitrofurans as veterinary drugs has been banned from intensive animal production in the European Union (EU) since 1993. The objective of the present study was to evaluate the accumulation and depletion of furaltadone and nifursol and their side-chain metabolites 5-methylmorpholino-3-amino-2-oxazolidinone (AMOZ) and 3,5-dinitrosalicylic acid hydrazide (DNSAH) in eggs after administration of therapeutic and subtherapeutic doses of the drugs to laying hens during three consecutive weeks. LC-MS/MS, with positive and negative electrospray ionization methods, was used for the determination of parent compounds and metabolites in yolk and egg white and was validated according to criteria established by Commission Decision 2002/657/EC. The decision limit (CCα) and the detection capability (CCβ) of the analytical methodology for metabolites were 0.1 and 0.5 μg/kg for AMOZ and 0.3 and 0.9 μg/kg for DNSAH, respectively. For the parent compounds, CCα and CCβ were 0.9 and 2.0 μg/kg for furaltadone and 1.3 and 3.1 μg/kg for nifursol, respectively. The data obtained show that the parent compounds are much less persistent than their side-chain metabolites in either yolk or egg white. Between the studied metabolites, AMOZ is the most persistent and could be detected in either yolk or egg white three weeks following withdrawal from treatment.     

Isolation and identification of sea buckthorn (Hippophae rhamnoides) phenolics with antioxidant activity and α-glucosidase inhibitory effect

This study was performed to evaluate the antioxidant and α-glucosidase inhibitory effects from the extract, fractions, and isolated compounds of sea buckthorn leaves. Six compounds, kaempferol-3-O-β-D-(6'-O-coumaryl) glycoside, 1-feruloyl-β-D-glucopyranoside, isorhamnetin-3-O-glucoside, quercetin 3-O-β-D-glucopyranoside, quercetin 3-O-β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside, and isorhamnetin-3-O-rutinoside, were isolated from sea buckthorn leaf extracts. The butanol fraction (EC(50) = 1.81 μg/mL) along with quercetin 3-O-β-D-glucopyranoside (EC(50) = 1.86 μg/mL) had a higher DPPH radical-scavenging activity and showed stronger reducing power (OD(700) = 1.83 and 1.78, respectively). The butanol fraction (477 mg GAE/g) contained the highest amount of phenolic compounds and also the most powerful α-glucosidase inhibitory effect (86%) at 5 μg/mL. The results indicate that sea buckthorn leaf extracts could potentially be used for food additives and the development of useful natural compounds.     

Simultaneous detection of residues of 25 β₂-agonists and 23 β-blockers in animal foods by high-performance liquid chromatography coupled with linear ion trap mass spectrometry

A sensitive method has been developed for the simultaneous determination of residues of 25 β?-agonists and 23 β-blockers in animal foods by high-performance liquid chromatography coupled with linear ion trap mass spectrometry (HPLC-LIT-MS). This method is based on a new procedure of hydrolysis and extraction by 5% trichloracetic acid, and then cleaned up by mixed strong cation exchange (MCX) cartridges coupled with a novelty cleanup step by methanol. Methanol and 0.1% formic acid were used as mobile phases for gradient elution, while a Supelco Ascentis Express Rp-Amide column was used for LC separation. ESI positive ion scan mode was used with consecutive reaction monitoring (CRM, MS3). Nine β?-agonists labeled by the deuterium isotope were used as internal standards for quantification. The linear ranges of 48 analytes were from 5 to 200 μg/L; the coefficient of correlation was not less than 0.995. Blank pork muscle, blank liver, and blank kidney were selected as representative matrix for spiked standard recovery test. The recoveries of each compound were in the range of 46.6-118.9%, and the relative standard deviations were in the range of 1.9-28.2%. Decision limits (CCα, α = 0.01) of 48 analytes in muscles, liver, and kidney samples ranged from 0.05 to 0.49 μg/kg, and the detection capability (CCβ, β = 0.05) ranged from 0.13 to 1.64 μg/kg. This method was successfully applied to 110 real animal origin food samples including meat, liver, and kidney of pig and chicken samples.     

SOME EFFECTS OF FERTILIZERS AND FARMYARD MANURE ON THE ORGANIC PHOSPHORUS IN SOILS

The amounts of P applied cumulatively to a neutral arable soil (pH 7.1–7. in 0.01M CaCl₂) at Rothamsted, as farmyard manure, alone or with superphosphate, which were converted to organic P in 100 years ranged from 18 to 44 μg P/g of soil (0–23 cm). Superphosphate alone (3300 kg P/ha) slightly lessened the total organic P in the soil. Neither farmyard manure nor super-phosphate significantly changed the amounts (38 to 42 μg P/g) of inositol penta- and hexaphosphate in these soils. In the surface layers (0–7.5 cm) of soils from permanent grassland at Rothamsted, superphosphate (3370kg P/ha) increased organic P by 134 μg P/g at pH 4.5 and 19 μg P/g at pH 6.5, about 6 and 1 per cent respectively of the P remaining from superphosphate applied cumulatively since 1858. In the sub-surface layers (7.5–23 cm) superphosphate increased organic P by 93 μg P/g at pH 4.5 and 62 μg P/g at pH 6.2, about 18 and 10 per cent respectively of the P remaining from superphosphate. The sum of inositol penta- and hexaphosphates accounted for 32 per cent at pH 4.5 and 21 per cent at pH 6.5 of the increases in organic P in the surface layers and 45 per cent and 26 per cent in the sub-surface layers at pH 4.5 and 6.5 respectively. Superphosphate (1260–2100 kg P/ha) applied intermittently or cumulatively increased total organic P by 19 to 52 μg P/g and inositol penta- and hexaphosphates by 13 to 17 μg P/g in acid tea soils (pH 3.2–3.4) from Georgia, U.S.S.R. Rock phosphate (510–1020kg P/ha) applied cumulatively had no effect on either the total organic P or the inositol P in acid tea soils (PH 3.6–3.7) from Ceylon.     

Soyasaponins resist extrusion cooking and are not degraded during gut passage in Atlantic salmon (Salmo salar L.)

The stability of soyasaponins in fish feed formulations was investigated. The level of soyasaponin Ab, Bb, Bc, Ba-2,3-dihydro-2,5-dihydroxy-6-methyl-4H-pyran-4-one (Ba-DDMP), Bb-DDMP, and Bc-DDMP was quantified in 15 samples of defatted soybean meal, two full fat soybean meals, and two soybean protein concentrates by reverse phase high-performance liquid chromatography. The total level of saponins in the 15 samples of commercial defatted soybean meal ranged from 4.8-6.8 micromol/g (5.1-7.0 g/kg). The two full fat meals contained 4.4 and 4.7 micromol/g whereas no saponins could be detected in the alcohol-extracted soybean protein concentrates. Fifteen batches of fish feed containing 20% defatted soybean meal were produced by twin-screw extrusion from the 15 different samples of defatted soybean meal. Extrusion did not reduce the total level of group B saponins, but the ratio between DDMP-conjugated group B saponins and non-DDMP-conjugated group B saponins was slightly reduced. A soybean-containing diet was fed to seawater adapted Atlantic salmon for 9 weeks. Yttrium oxide was included in the feed as an inert marker in order to estimate the disappearance of saponins during gut passage. High levels of intact non-DDMP-conjugated group B soyasaponins were found in feces whereas only low levels of DDMP-conjugated saponins could be detected. The overall disappearance of saponins was close to zero, and the concentration of intact saponins in dry feces reached levels several fold higher than dietary levels. The present work demonstrates that non-DDMP-conjugated group B soyasaponins resist extrusion cooking and remain intact during gut passage in Atlantic salmon. The latter is contrary to earlier findings in endothermic animals.