Influence of phosphorus nutrition on growth and metabolism of Duo grass (Duo festulolium)

Use of suitable plants that can extract and concentrate excess P from contaminated soil serves as an attractive method of phytoremediation. Plants vary in their potential to assimilate different organic and inorganic P-substrates. In this study, the response of Duo grass (Duo festulolium) to variable rates of soil-applied potassium dihydrogen phosphate (KH₂PO₄) on biomass yield and P uptake were studied. Duo grown for 5 weeks in soil with 2.5, 5 and 7.5 g KH₂PO₄ kg^?1 soil showed a significantly higher biomass and shoot P content of 8.3, 11.4 and 12.3 g P kg^?1 dry weight respectively compared to plants that received no soil added P. Also, the ability of Duo to metabolize different forms of P-substrates was determined by growing them in sterile Hoagland's agar media with different organic and inorganic P-substrates, viz. KH₂PO₄, glucose-1-phosphate (G1P), inositiol hexaphosphate (IHP), adenosine triphosphate (ATP) and adenosine monophosphate (AMP) for 2 weeks. Plants on agar media with different P-substrates also showed enhanced biomass yield and shoot P relative to no P control and the P uptake was in the order of ATP > KH₂PO₄ > G1P > IHP = AMP > no P control. The activities of both phytase (E.C.3.1.3.26) and acid phosphatases (E.C.3.1.3.2) were higher in all the P received plants than the control. Duo grass is capable of extracting P from the soil and also from the agar media and thus it can serve as possible candidate for phytoextraction of high P-soil.     

l-Lactic acid production from liquefied cassava starch by thermotolerant Rhizopus microsporus: Characterization and optimization

The thermotolerant Rhizopus microsporus DMKU 33 capable of producing l-lactic acid from liquefied cassava starch was isolated and characterized for its phylogenetic relationship and growth temperature and pH ranges. The concentrations of (NH₄)₂SO_4, KH₂PO₄, MgSO₄ and ZnSO₄·7H₂O in the fermentation medium was optimized for lactic acid production from liquefied cassava starch by Rhizopus microsporus DMKU 33 in shake-flasks at 40 °C. The fermentation was then studied in a stirred-tank bioreactor with aeration at 0.75 vvm and agitation at 200 rpm, achieving the highest lactic acid production of 84 g/L with a yield of 0.84 g/g at pH 5.5 in 3 days. Lactic acid production was further increased to 105–118 g/L with a yield of 0.93 g/g and productivity of 1.25 g/L/h in fed-batch fermentation. R. microsporus DMKU 33 is thus advantageous to use in simultaneous saccharification and fermentation for l-lactic acid production from low-cost starchy substrates.     

Relative bioavailabilities of organic zinc sources with different chelation strengths for broilers fed diets with low or high phytate content

A six-day experiment was conducted to estimate the relative bioavailability values (RBV) of zinc (Zn) in three organic sources (oZn) with different chelation strengths compared to inorganic ZnSO₄ (iZn) for broilers fed a low or high phytate diet. A total of 1080, one-d-old male broiler chicks were randomly assigned to one of 18 dietary treatments (six replicates cages of ten chicks per cage) in a completely randomized design involving a 2 × 2 × 4 factorial arrangement with two levels of added phytate (0 or 10 g phytate as sodium phytate/kg), two levels of added Zn (30 or 60 mg/kg) and four Zn sources (iZn and three oZn sources) plus one low and one high phytate control treatments without Zn addition. The three oZn sources consisted of (1) Zn amino acid with weak chelation strength (ZnAA-L, formation quotient Q_f = 6.6, containing 119 g Zn/kg), (2) Zn proteinate with moderate chelation strength (ZnPRO-M, Q_f = 30.7, containing 133 g Zn/kg) or (3) Zn proteinate with strong chelation strength (ZnPRO-H, Q_f = 944.0, containing 186 g Zn/kg). Chicks were harvested at 6 days of age and pancreas metallothionein (MT) mRNA expression was used to estimate Zn RBV. Pancreas MT mRNA expression increased (P<0.01) as dietary Zn level increased. Chicks fed high phytate diets had lower (P<0.05) MT mRNA expression than chicks fed low phytate diets. Based on multiple linear regression slope ratios with ZnSO₄ set at 1.00, the RBV of ZnAA-L, ZnPRO-M and ZnPRO-H were 1.01, 1.28 and 0.70, respectively, for low phytate diets, and 1.05, 1.39 and 0.92, respectively, for high phytate diets. The slope for the oZn source with moderate chelation strength differed (P<0.05) from iZn and the other two oZn sources. The RBV of ZnAA-L, ZnPRO-M and ZnPRO-H under the high phytate diet increased by 0.04, 0.11 and 0.22, respectively, compared to those under the low phytate diet. Results indicate that the oZn sources with moderate or strong chelation strength offer partial or complete resistance to interference from high dietary phytate during digestion; and the oZn with moderate chelation strength had a greater RBV with both low and high phytate diets than iZn or oZn sources with weak or strong chelation strength.     

Nutritional characterization of Mucuna pruriens: 3. Effect of replacing soybean meal with Mucuna on intake,digestibility, N balance and microbial protein synthesis in sheep

Mucuna pruriens seeds have relatively high crude protein (CP) concentrations, but little is known about their potential to replace commonly used CP supplements in ruminant rations. The aim of this experiment was to determine effects of replacing soybean meal (SB) with Mucuna on the performance of lambs. Forty Rambouillet lambs (33.2 ± 5.73 kg) fed a basal diet of maize grain, cottonseed hulls and urea were randomly assigned to one of four supplements formulated by substituting 0 (SB), 330 (Lo), 670 (Med) or 1000 g/kg (Hi) of soybean meal with rolled Mucuna seeds. Lambs were housed individually in metabolic crates and allowed ad libitum access to isocaloric (metabolizable energy=11.7 MJ/kg dry matter, DM) and isonitrogenous (CP = 146 g/kg, DM) diets for 14 d of adaptation and 7 d of total fecal collection. Fecal egg counts and coccidian oocyst scores were determined on d 14. Dry matter intake (1.7 kg/d versus 1.5 kg/d; P<0.05), CP digestibility (774 g/kg versus 714 g/kg DM; P<0.05) and N retention (28.0 g/d versus 20.4 g/d; P<0.01) were higher and amylase-pretreated neutral detergent fiber digestibility (617 g/kg versus 686 g/kg DM) was lower (P<0.05) in sheep fed SB versus Mucuna diets. However, supplementary protein source did not affect rumen pH, blood urea N or glucose concentration, or fecal egg counts. Increasing the level of Mucuna supplementation increased (P<0.05) level and efficiency of microbial protein synthesis, ruminal fluid acidity, total volatile fatty acid concentration, decreased (P<0.05) coccidian oocyst scores, and tended (P<0.10) to increase N retention. Therefore, SB is a better supplement than Mucuna to support performance of lambs. Nevertheless, Mucuna seeds are a promising CP supplement for situations where cost or availability precludes use of SB in ruminant rations.     

Comparative studies on dry matter intake,digestibility and nitrogen metabolism between Thai native (TN) and Anglo Nubian × TN bucks

Metabolic experiments were conducted for each Thai native (TN) and Anglo Nubian–TN 50% (AN × TN) bucks, 26.5–38 kg BW and 35–52 months of age, by assigning the following concentrate diets, which had four levels of CP concentrations, T1: 14%, T2: 21%, T3: 28% and T4: 35%, on a DM basis. The animals were fed the concentrates at a rate of 1% of their BW and had ad libitum access to Paspalum plicatulum hay. As the CP level increased, digestibility of CP, N excretion in urine and N retention increased (P < 0.01). No significant differences of DM digestibility among the CP levels of diet and between the breeds of bucks were observed. The AN × TN had higher DM intake than TN (P < 0.05, 40.9 g/(kg BW^0.75 day) versus 36.0 g/(kg BW^0.75 day)), thus N intake of AN × TN was higher than that of TN (P < 0.01, 0.99 g/(kg BW^0.75 day) versus 0.90 g/(kg BW^0.75 day)). Digestibility of CP and blood urea N concentrations of TN were higher than those of AN × TN (P < 0.05, 69.8% versus 64.0% and 32.2 mg/dl versus 26.7 mg/dl, respectively). The AN × TN tended to retain more N than TN in the higher CP feeding condition of T3 (0.19 g/(kg BW^0.75 day) versus 0.11 g/(kg BW^0.75 day)) and T4 (0.25 g/(kg BW^0.75 day) versus 0.17 g/(kg BW^0.75 day)). Digestibility of NDF and ADF, and TDN of TN were higher than those of AN × TN (P < 0.05, 61.1% versus 55.9%, 49.6% versus 43.5% and 65.8% versus 62.4%, respectively). The overall mean DE was 109 kcal/(kg BW^0.75 day), and no significant difference of DE among CP levels nor breeds of bucks was observed. Thai native was superior to AN × TN in digestibility of N and fiber fractions at the maintenance level of DE, which might have overcome inferiority of TN to AN × TN in amount of N and energy intake caused by the lower DMI. It is likely that TN goats are well adapted to fodder shortage condition due to their efficient utilization of nutrients.     

The interaction between lactose level and crude protein concentration on piglet post-weaning performance,nitrogen metabolism,selected faecal microbial populations and faecal volatile fatty acid concentrations

A performance study and a nitrogen balance study (2×3 factorial) were conducted to investigate the interaction between lactose level (215 and 125 g/kg) (lactofeed 70; 860 g whey permeate/kg, 140 g soya bean meal/kg, Volac International, UK) and crude protein (CP) concentration (160, 185 and 210 g/kg) on post-weaning piglet performance, nitrogen metabolism, faecal microbiology and faecal volatile fatty acid concentrations. In the performance trial, 252 piglets (7.6 kg; 33 days of age) were assigned to one of six dietary treatments following a 12-day period on a commercial creep diet (17 MJ/kg DE, 16 g lysine/kg). The experimental diets were fed for 28 days (days 12–40) and were formulated to have identical digestible energy (15 MJ/kg) and total lysine (14.5 g/kg) contents. In the N balance experiment, 24 boars (20 kg live weight) were offered the same diets as in the performance trial. Faecal samples were collected for selected microbial populations. There was an interaction (P<0.05) between lactose and CP concentration in daily gain (ADG) and daily feed intake (ADFI) (P<0.01) during the weaner period (days 12–40). At the high lactose level there was a linear increase in ADG and ADFI with increasing CP. However, at the low lactose level there was no increase in ADG or ADFI above the medium CP. Pigs offered 215 g lactose/kg had a higher dry matter (P<0.001), organic matter (P<0.001), energy (P<0.001), nitrogen (P<0.01) and neutral detergent fibre (P<0.05) coefficient of total tract apparent digestibility compared to pigs offered 125 g lactose/kg. There was an interaction between lactose and CP concentration for nitrogen intake (NI) (P<0.05), urine pH (P<0.05) and selected faecal microbial populations. At the high CP level, pigs offered diets containing 215 g lactose/kg had a higher NI and a lower urine pH than pigs offered 125 g lactose/kg (P<0.05). However, the inclusion of lactose had no significant effect on either NI or urine pH at the low or medium CP concentration. At the low lactose level there was a linear increase in faecal E. coli population and a linear decrease in faecal Lactobacilli population with increasing CP. However at high lactose levels CP concentration had no effect on either E. coli or Lactobacilli populations. Pigs offered 215 g lactose/kg had a significantly higher Bifidobacteria population compared to pigs offered 125 g lactose/kg. There was a linear decrease in Bifidobacteria population as CP increased. In conclusion, at the high lactose level there was a linear increase in ADG and ADFI with increasing CP concentrations. There was no increase in these parameters above 185 g CP/kg at the low lactose level.     

The effect of dietary crude protein level on intake,growth, protein retention and utilization of growing male Saanen kids

The effects of increasing dietary levels of crude protein on growth, feed intake, feed efficiency and composition of gain in male Saanen kids were studied. Four groups of four kids each initially weighing 12.1±0.18 kg and having a weight gain of 195±16 g/d were penned individually and fed for 73 to 124 days up to 25 kg of BW. They were fed chaffed wheat straw (45 g CP/kg DM) which had been molasses sprayed (10%) and pelleted concentrates containing 8.7, 11.7, 14.4 and 17.6% crude protein (CP) on DM basis, the ratio of straw to concentrate being 1:5. Kids were fed controlled to satiation in which case small amounts of the feeds were offered about five times a day as long as the kids wanted to eat. Retention of protein, fat and energy were calculated from their initial and final concentrations in the empty body homogenates of the slaughtered kids. With increasing CP level in the diet, feed intake increased from 448 to 608 g DM/d, weight gain from 94 to 181 g/d, retention of protein from 9.7 to 27.8 g/d, retention of fat from 9.6 to 19.1 g/d and feed efficiency improved from 4.79 to 3.39 kg DM/kg weight gain. Protein composition of gain increased from 103 to 154 g/kg BWG while fat (103–105 g/kg BWG) remained constant. Regression analyses showed that BWG can be optimized at 136 g CP/kg DM and protein retention at 180 g CP/kg DM, whereas, dietary nitrogen was utilized most efficiently at 120 g CP/kg DM. Extrapolated from the regression equations, maintenance N requirement of the kids at zero N-retention and at zero BWG were 0.38 and 0.16 g N/kg W^0.75, respectively. Recommended dietary CP concentrations and maintenance N requirements depend on the traits desired.     

Efficient production of α-arbutin by whole-cell biocatalysis using immobilized hydroquinone as a glucosyl acceptor

The aim of the present study is to develop an efficient and cost-effective method for α-arbutin production by using whole-cell of Xanthomonas maltophilia BT-112 as a biocatalyst. Hydroquinone (HQ), substrate for the bioconversion as glucosyl acceptor, was immobilized on H107 macroporous resin to reduce its toxic effect on the cells, and the optimal reaction conditions for α-arbutin synthesis were investigated. When 350 g/L H107 resin (254.5 mM HQ) and 20 g/L (4.2 U/g) of cells were shaken in 10 mL Na₂HPO₄–KH₂PO₄ buffer (50 mM, pH 6.5) containing 509 mM sucrose at 35 °C with 150 rpm for 48 h, the final yield of α-arbutin reached 65.9 g/L with a conversion yield of 95.2% based on the amount of HQ supplied. The α-arbutin production was 202% higher than that of the control (free HQ) and the cells maintained its full activity for almost six consecutive batch reactions, indicating a potential for reducing production costs. Additionally, the product was one-step isolated and identified as α-arbutin by ¹³C NMR and ¹H NMR analysis. In conclusion, the combination of whole cells and immobilized hydroquinone (IMHQ) is a promising approach for economical and industrial-scale production of α-arbutin.     

Effect of modified MRS medium on production and purification of antimicrobial peptide ST4SA produced by Enterococcus mundtii

Highest antimicrobial activity of peptide ST4SA (51,200 AU/mL) was recorded after 14 h of growth in MRS broth with optimal production at pH 6.0 or 6.5. Growth of strain ST4SA in the presence of tryptone, yeast extract, or a combination of the two, yielded 102,400 AU/mL. An increase in production of peptide ST4SA to 102,400 AU/mL was recorded in the presence of 20.0 g/L fructose, but decreased to 25,600 AU/mL in the presence of lactose (20.0 g/L) or mannose (20.0 g/L) as sole carbon source. Lower activity (25,600 AU/mL) was recorded when 2.0 g/L K₂HPO₄ was replaced by 2.0 g/L KH₂PO₄ in MRS broth. An increase of K₂HPO₄ to 10.0 g/L and 20.0 g/L resulted in higher activity (102,400 AU/mL). Addition of glycerol to MRS broth had a negative effect on peptide ST4SA production. Production of peptide ST4SA required the presence of magnesium sulphate, manganese sulphate and 5.0 g/L sodium acetate. Exclusion of tri-ammonium citrate from the medium resulted in reduction of activity to 3,200 AU/mL. Maximum activity (102,400 AU/mL) was recorded in MRS supplemented with 1.0 ppm Vit. C, DL-6,8-thioctic acid or thiamine, respectively. Growth of Listeria ivanovii susbp. ivanovii ATCC 19119 in the presence of peptide ST4SA (12,800 AU/mL) resulted in 99% cell lysis after 18 h. Improved production of peptide ST4SA was recorded in MRS broth (Biolab) pre-treated with Amberlite XAD-1180. Precipitation with ammonium sulphate, followed by gel filtration chromatography, yielded the highest level of peptide ST4SA. This paper describes the partially deproteination of growth medium to facilitate peptide ST4SA purification.     

Microbial protein synthesis in growing-finishing bulls estimated from the urinary excretion of purine derivatives

In the frame of a feeding experiment with three periods, balance trials were carried out with 18 double-muscled Belgian White–blue bulls, allocated to one of four feeding regimes. The ration consisted of maize silage and concentrate in the ratio of 35:65 on DM basis. Six concentrates were formulated to supply three levels of protein and energy. The three periods corresponded to distinct live weight intervals of 360–460, 460–570 and 570–680 kg. In the first feeding regime, low protein and intermediate energy level were given during the whole trial; in the second regime, protein level decreased at a constant intermediate energy level; in the third regime, the energy level increased at a constant high protein level; in the fourth regime, protein level decreased simultaneously with an increase in the energy level. Total daily intake varied from 6.2 to 9.7 kg for dry matter (DM), from 65 to 106 MJ for metabolisable energy (ME) and from 719 to 1326 g for crude protein (CP) (50 animal observations). At the end of each period, the excretion of the purine derivatives (PD), allantoin and uric acid, was measured after total urine collection during 4 days to estimate microbial nitrogen supply to the duodenum (MN_PD). The effect of the intake of DM, organic matter (OM), digestible OM (DOM), digestible carbohydrates (DCHO), total digestible nutrients (TDN), rumen fermentable OM (FOM), metabolisable energy (ME), fermentable ME (FME), CP, digestible CP (DCP) and rumen degradable protein (RDP) on PD and MN_PD was examined. Further, the relationship of MN_PD to MN, calculated according to different systems, was examined. The amount (mean±SD) of allantoin excreted in urine was 147±23 mmol day⁻¹ and of uric acid 11±3 mmol day⁻¹. The MN_PD amounted to 97±21 g day⁻¹, varying from 57 to 154 g day⁻¹, and significantly increased with all measures of nutrient intake. The correlation coefficients, ranging from 0.45 for DCP to 0.57 for DOM and FME, were, however, not significantly different. MN_PD showed a larger variation and was on average lower than the potential MN values calculated from the intake of FOM (108±13 g day⁻¹), DCHO (141±16 g day⁻¹) and FME (109±13 g day⁻¹), similar to that calculated from the intake of RDP (99±14 g day⁻¹) and higher than the MN value calculated from the intake of TDN (84±11 g day⁻¹). The correlations of MN_PD to the calculated MN values ranged from 0.47 for MN_FOM to 0.59 for MN_FME, but were not significantly different.     

A pharmaceutical preparation of Salvia miltiorrhiza protects cardiac myocytes from tumor necrosis factor-induced apoptosis and reduces angiotensin II-stimulated collagen synthesis in fibroblasts

Salvia miltiorrhiza is a medicinal herb commonly used in traditional Chinese medicine for the prevention and treatment of cardiovascular disease. This study investigated the effects of Cardiotonic Pill (CP), a pharmaceutical preparation of Salvia miltiorrhiza, on cardiac myocytes and fibroblasts with respect to the viability, proliferation, and collagen synthesis in these cells under various conditions. A cardiac myocyte line, H9c2, and primarily cultured fibroblasts from rat hearts were incubated with CP over a broad concentration range (50–800 μg/ml) under normal cultures, conditions of ischemia (serum-free culture), and stimulation by angiotensin II (AII, 100 nM), hydrogen peroxide (H₂O₂, 50–200 μM), or tumor necrosis factor α (TNFα, 40 ng/ml) for 24–48 h. Cell growth, apoptosis, DNA and collagen synthesis, and expression of relevant genes were assessed via cell number study, morphological examination, Annexin-V staining, flow-cytometry, [³H]-thymidine or [³H]-proline incorporation assay, and Western blotting analysis. It was found that (1) at therapeutic (50 μg/ml) and double therapeutic (100 μg/ml) concentrations, CP did not significantly affect normal DNA synthesis and cell growth in these cardiac cells, while at higher (over 4-fold therapeutic) concentrations (200–800 μg/ml), CP decreased DNA synthesis and cell growth and increased cell death; (2) CP treatment (50 μg/ml) significantly inhibited TNFα-induced apoptosis in myocytes, with 12.3±1.46% cells being apoptosis in CP treatment group and 37.0±7.34% in the control (p<0.01), and simultaneously, expression of activated (phosphorylated) Akt protein was increased by about 2 folds in the CP-treated cells; and (3) in cultured fibroblasts, CP significantly reduced AII-induced collagen synthesis in a concentration-dependent manner (by ～50% and ～90% reduction of AII-induced collagen synthesis at 50 and 100 μg/ml, respectively). Thus, Salvia miltiorrhiza preparation CP is physiologically active on cardiac cells. The actions by CP to reduce apoptotic damage in myocytes and collagen synthesis in fibroblasts may help to preserve the heart function and reduce heart failure risk. The actions by CP to inhibit DNA synthesis and cell growth, which occurred at over therapeutic doses, may weaken the ability of heart repair. Further studies are needed to identify the chemical compounds in this herbal product that are responsible for these observed physiological effects.     

Effect of feeding schedule on apparent energy and amino acid digestibility by growing pigs

An experiment was conducted to investigate the effects of feeding schedule on dry matter (DM), gross energy (GE), crude protein (CP), and amino acid (AA) digestibility by growing pigs. Six growing castrates (initial body weight: 40.5 ± 2.25 kg) were equipped with a T-cannula in the distal ileum and allotted to a repeated 3 × 3 Latin square design with three animals and three periods in each square. A maize–soybean meal based diet (181 g CP/kg) was formulated and fed to the pigs in three different feeding schedules; i.e., one meal per day, two meals per day, or free access to feed. Pigs fed one or two meals per day were allotted a daily amount of feed equivalent to three times the maintenance energy requirement. Fecal and ileal samples were collected from all pigs in each period. The coefficients of ileal apparent digestibility (CIAD) for DM, GE, CP, and AA were calculated as were the coefficients of total tract apparent digestibility (CTTAD) of DM and GE. Results of the experiment showed that there were no differences in the CIAD for DM, GE, or for any of the AA among the three feeding schedules. In contrast, pigs given free access to the diet had lower (P<0.002) CTTAD for DM and GE compared with pigs fed a restricted amount of feed either once or twice daily. Therefore, the concentration of digestible energy in the diet was lower if calculated from pigs given free access to the diet compared with pigs fed the diet once or twice daily (14.38 MJ/kg versus 14.83 MJ/kg and 15.13 MJ/kg). It is concluded that the feeding schedule does not influence the CIAD for DM, GE, CP, or AA by pigs. Therefore, any of the three feeding schedules that were used in the current experiment may be used in experiments aimed at measuring CIAD in pigs. However, if the CTTAD of DM and GE are measured, it is recommended that pigs are given free access to feed.     

The effect of variety on maize grain and crop residue yield and nutritive value of the stover

Eight varieties of maize, namely Awassa 511 (A511), Birkata, composite of best families (CBF), Dendane, drought tolerant population (DTP1), Guto, Katumani and Keroshet planted in 1995 cropping season were compared for grain and crop residue yield and nutritive value of the stover. Significant differences (p < 0.05) were observed in grain and crop residue yields of the varieties. Grain yield was highest (6.99 t ha⁻¹) in DTP1 and lowest (2.21 t ha⁻¹) in Keroshet. Stover and total crop residue yields were significantly higher (p < 0.05) in A511 than in CBF and Dendane. The stem was the largest portion in the stover varying from 31% in Dendane to 50% in A511. The stover of Birkata showed a high crude protein (CP) and low ash and fibre contents compared to the other varieties. The CP content varied from 28 g kg DM⁻¹ (Guto) to 61 g kg DM⁻¹ (Birkata). Crop residue yield and quality parameters, except CP content of the stover, were not negatively correlated with grain yield. The potential utility index varied from 61% (Keroshet) to 74% (CBF). The varieties with higher potential utility indices, except Guto, also had relatively higher solubility, 48 h DM degradability and effective degradability compared to the other varieties. The overall results showed evidence of varietal differences in grain and stover yield and stover quality and indicated the possibility of selecting for maize varieties that combine high grain yield and desirable stover characteristics.     

Statistical optimization of critical medium components for lipase production from Yarrowia lipolytica (MTCC 35)

Statistical optimization is an effective technique for the investigation of complex processes with minimal number of experimental runs. In this study, statistical approach was used to study the optimization of media components for lipase production from Yarrowia lipolytica MTCC 35. Mahua cake, glucose, MnCl₂ and KH₂PO₄ were screened to be the most significant variables among the nine medium variables that were tested to determine influence on lipase production by Plackett–Burman design. Central Composite Design was used for further optimization of these screened variables for enhanced lipase production. The determination coefficient (R²) value of 0.922 showed that the regression models adequately explain the data variation and represent the actual relationships between the variables and response. The optimum values of investigated variables for the maximum lipase production were 6.0% Mahua cake, 2.0% glucose, 0.2% MnCl₂ and 0.2% KH₂PO₄. The maximum lipase production (9.40 U mL^?1) was obtained under optimal condition.     

Optimization of date syrup for enhancement of the production of citric acid using immobilized cells of Aspergillus niger

Date syrup as an economical source of carbohydrates and immobilized Aspergillus niger J4, which was entrapped in calcium alginate pellets, were employed for enhancing the production of citric acid. Maximum production was achieved by pre-treating date syrup with 1.5% tricalcium phosphate to remove heavy metals. The production of citric acid using a pretreated medium was 38.87% higher than an untreated one that consumed sugar. The appropriate presence of nitrogen, phosphate and magnesium appeared to be important in order for citric acid to accumulate. The production of citric acid and the consumed sugar was higher when using 0.1% ammonium nitrate as the best source of nitrogen. The production of citric acid increased significantly when 0.1 g/l of KH₂PO₄ was added to the medium of date syrup. The addition of magnesium sulfate at the rate of 0.20 g/l had a stimulating effect on the production of citric acid. Maximum production of citric acid was obtained when calcium chloride was absent. One of the most important benefits of immobilized cells is their ability and stability to produce citric acid under a repeated batch culture. Over four repeated batches, the production of citric acid production was maintained for 24 days when each cycle continued for 144 h. The results obtained in the repeated batch cultivation using date syrup confirmed that date syrup could be used as a medium for the industrial production of citric acid.     

Evaluation of fungal contamination and mycotoxin production in maize silage

Agricultural activities involve daily use of maize silage as feed for livestock, which can be contaminated by mycotoxigenic molds. To evaluate fungal contamination, and the production of mycotoxins in maize silage we propose a multi-disciplinary approach utilizing PCR methods with genes of the aflatoxin (ver-1, omt-1 and apa-2), fumonisin (FUM1) and trichothecene (TRI6) biosynthesis pathways. To detect Aspergillus fumigatus, a 26S/intergenic spacer region of the rDNA complex was amplified. These specific PCR assays allowed three major groups of toxigenic fungi-like aflatoxin-producing Aspergilli, fumonisin and trichothecene-producing Fusaria, and the ubiquitous mold A. fumigatus, to be targeted. A multimycotoxin method is also proposed to simultaneously quantify seven mycotoxins (i.e., aflatoxin B₁, citrinin, deoxynivalenol, fumonisin B₁, gliotoxin, ochratoxin A, zearalenone) in maize silage by high-performance liquid chromatography coupled to mass spectrometry (HPLC–MS). These microbiological and analytical tools revealed three potentially toxigenic groups of fungi and A. fumigatus grown from mature maize silage (11 month old) that was collected in Normandy (France) and the mycotoxins aflatoxin B₁ (7.0–51.3 μg/kg), citrinin (10.1–14.2 μg/kg), deoxynivalenol (128.0–181.0 μg/kg) and gliotoxin (6.6–11.9 μg/kg). Results indicate that the combination of PCR and HPLC–MS can be used to assess fungal quality of maize silages.     

Effects of urea and aqueous ammonia treatment on the composition and nutritive value of maize residues

Two trials were conducted to investigate changes in chemical composition and nutritive value of maize residues treated with urea and aqueous ammonia. In Experiment I, maize stalks, husks and cobs were chopped into pieces of approximately 1 cm length. Aqueous NH₃ (3%) and feed grade urea equivalent to 3% NH₃ were sprayed into 100 g duplicate samples on DM basis, thoroughly mixed, stored in plastic containers and kept for 3 week at ambient temperature (25–27 °C). Proximate composition and in vitro dry matter digestibility of test materials were determined. In Experiment II, 100 kg batches of the maize residues were chopped and treated as in Experiment I but stored in 205 l drums lined with black polythene sheet. Feed intake and nutrient digestibility of treated materials were evaluated with 18 mature WAD sheep averaging 19.8 kg BW. Treatment improved (P < 0.05) intake and digestion coefficients for N, DM, NDF, ADF and OM but there were no differences (P > 0.05) between aqueous NH₃ and urea treatments. Feed grade urea or the equivalent weight of fertilizer grade urea can be used to improve the nutritional value of maize residues for small ruminant feeding during off season periods.     

Mycophenolic acid production in solid-state fermentation using a packed-bed bioreactor

Mycophenolic acid (MPA) was produced from Penicillium brevicompactum by solid-state fermentation (SSF) using pearl barley, and submerged fermentation (SmF) using mannitol. It was found that SSF was superior to SmF in terms of MPA concentration (1219 mg/L vs. 60 mg/L after 144 h fermentation), and the product yields were 6.1 mg/g pearl barley for SSF and 1.2 mg/g mannitol for SmF. The volumetric productivities were 8.5 and 0.42 mg/L h for SSF and SmF, respectively.The optimum solid substrate of SSF for MPA production was pearl barley, producing 5470 mg/kg compared with wheat bran (1601 mg/kg), oat (3717 mg/kg) and rice (2597 mg/kg). The optimum moisture content, incubation time and inoculum concentrations were 70%, 144 h and 6%, respectively. Neither the addition of mannitol or (NH4)₂HPO₄ nor adjustment of media pH within the range of 3–7 significantly enhanced MPA production.MPA production by SSF using a packed-bed bioreactor was performed and an increased maximum production of MPA 6.9 mg/g was achieved at 168 h incubation time. The higher volumetric productivity and concentrations makes SSF an attractive alternative to SmF for MPA production.     

L-Citrulline acts as potential hypothermic agent to afford thermotolerance in chicks

Recently we demonstrated that L-citrulline (L-Cit) causes hypothermia in chicks. However, the question of how L-Cit mediates hypothermia remained elusive. Thus, the objective of this study was to examine some possible factors in the process of L-Cit-mediated hypothermia and to confirm whether L-Cit can also afford thermotolerance in young chicks. Chicks were subjected to oral administration of L-Cit along with intraperitoneal injection of a nitric oxide synthase (NOS) inhibitor, N^G-nitro-l-arginine methyl ester HCl (L-NAME), to examine the involvement of NO in the process of hypothermia. Food intake and plasma metabolites were also analyzed after oral administration of L-Cit in chicks. To examine thermotolerance, chicks were orally administered with a single dose of L-Cit (15 mmol/10 ml/kg body weight) or the same dose twice within a short interval of 1 h (dual oral administration) before the exposure to high ambient temperature (35 ± 1 °C) for 180 min. Although the rectal temperature was reduced following administration of L-Cit, L-NAME caused a greater reduction. L-NAME reduced total NO₂ and NO₃ (NOx) in plasma, which confirmed its inhibitory effect on NO. A single oral administration of L-Cit mediated a persistent state of hypothermia for the 300 min of the study without affecting food intake. It was further found that plasma glucose was significantly lower in L-Cit-treated chicks. Dual oral administration of L-Cit, but not a single oral administration, afforded thermotolerance without a significant change in plasma NOx in chicks. In conclusion, our results suggest that L-Cit-mediated hypothermia and thermotolerance may not be involved in NO production. L-Cit-mediated thermotolerance further suggests that L-Cit may serve as an important nutritional supplement that could help in coping with summer heat.     

Statistical media optimization for growth and PHB production from methanol by a methylotrophic bacterium

Media components were optimized by statistical design for cell growth and PHB production of Methylobacterium extorquens DSMZ 1340. Four important components of growth media were optimized by central composite design. The growth increased from an OD = 1.35 for Choi medium as control to an OD = 2.15 for optimal medium. Then media components for PHB production were optimized. Optimization of five important factors was conducted by response surface method. The optimal composition of PHB production medium was found to be at 7.8 (g/L) Na₂HPO₄ · 12H₂O, and surprisingly at zero concentration of (NH₄)₂SO₄, KH₂PO₄, MgSO₄ and MnSO₄. The PHB production was found to be 2.95 (g/L) at this medium. RSM results indicated that a deficiency of nitrogen and magnesium is crucial for PHB accumulation in this microorganism. Also, PHB production was carried out in a 5 L fermentor at the optimum condition which resulted in 9.5 g/L PHB and 15.4 g/L cell dry weight with 62.3% polymer content.