Cattle and porcine Biol sampling

Two Sistemas Biobolsa^® (IRRI of Mexico) of 6000 and 12 000 L were installed in the agricultural sector of the Technological Institute of the Tlaxcala Altiplano (ITAT), Mexico. The bioles were sampled during May and June. Samples were taken at the exit of the Sistemas Biobolsa fed with cattle and porcine manure in a 1: 3 and 1: 2 (fresh basis) respectively. We collected samples with biodigester systems in good physical conditions and proper functioning. Then, to homogenize the liquid within the biobolsa, we hand rubbed each digester for 5 min. We placed homogeneous portions of biol with a collecting cane in clean plastic containers of 600 mL and screw cap; we took one sample from the digester located in the effluent stream. To avoid collecting samples at noon, when heat causes the dilation of the biobolsa and obstructs its homogenisation, we did sampling between 10:00 and 11:00 h. The collected samples were stored at 4 °C for up to 4 h before performing analysis.

Temperature, pH, electrical conductivity (EC), total dissolved solids (TDS), salinity and dissolved oxygen (DO)

With a pH meter (HANNA HI-255) we determined: temperature, pH, EC, TDS, and salinity (NaCl) in bioles of cattle and porcine manure. The procedures were performed according to the standards ^{NMX-AA-007-SCFI-2000}, NMX-AA008-SCFI-2000, ^{NMX-AA-034-SCFI-2001}, for temperature, pH, and total dissolved solids respectively. We made calibration curves for pH with samples from three points we calibrated conductivity with a standard solution of 1413 μScm^-1 (HI 7031), the SDT with a solution of TDS (HI-7032) and salinity with a standard solution (HI-7073), all of them were HANNA brand.

Dissolved oxygen (DO) and oxidation-reduction potential (ORP)

To determine OD and ORP we used glass bottles for BOD and a multiparameter meter (HACH, model Q40d with probes for dissolved oxygen) was utilized to evaluate LDO and ORP. These electrodes were calibrated with ambient oxygen and a ORP (HI-7022 and HANNA) standard solution.

Water percentage, total solids (TS), volatile solids (SV) and ashes

Humidity, ST, SV and ashes were calculated by weight difference after keeping the samples at 105 °C and calcined at 550 °C; the analyses were performed in duplicate. The formulas to calculate the water content, ST, SV and ash, expressed as percentages, were:

where a: mass in grams of wet biol; b: mass in grams of biol dehydrated at 105 °C ±5 °C; c: mass in grams of biol calcined at 550 ºC ± 5 ºC.

Analysis of macronutrients: phosphorus as P ₂ O ₅ , PO ₄ , ammonia nitrogen (NH ₃ - H) and potassium

Biol was centrifuged at 5000 rpm for 15 min in a centrifuge Hermle Z323; the supernatant was used for quantification of P, as P₂O₅ and PO₄^-3, and NH3-H. Analyses were performed in a nutrient analyzer HANNA HI-83215; K was quantified in an atomic absorption spectrophotometer GBC AA 904.

Elemental analysis of micronutrients

Elemental analyses were carried out with an atomic absorption spectrophotometer GBC AA 904. For the analysis of each element we prepared calibration curves at three points with standards of known concentrations. We used hollow cathode lamps of sodium, zinc, magnesium, copper and calcium for each element. For each element we used the air-acetylene burner, except for calcium, in which the burner N₂O acetylene was used. The followed was according to the ^{NMX-AA-051-SCFI-2001} standard. The differences between the type of biol used were evaluated by One Factor Analysis of Variance for all physical and chemical parameters, and ANOVA at an Excel^® factor.

Temperature, pH, CE, SDT and salinity

The average temperature during two months in cattle and porcine bioles was 22.4 and 23.13 ºC ± 1.7 ºC; these temperatures depended on the sampling time. This temperature range promotes mainly the reproduction of mesophilic microorganisms (20-40 °C); low temperature affects the rate of production of methane. Although fermentation was not carried out in thermophilic conditions, considered optimal (^{Yokoyama et al., 2007}; ^{Nagy and Wopera, 2012}), methane was generated up to 60 %.

The variation of pH during the 45 d remained within the optimal range, confirming the proper operation of the digester. The pH of cattle and porcine bioles is similar and shows some relationship with those documented by ^{Quipuzco et al. (2011)}, with neutral or slightly alkaline range. These values showed that bioles might be applied to all crops, including fruit trees. The pH and temperature have a direct effect on the rate of methane generation (^{González and Longoria, 2005}; ^{Amon et al., 2006}). Therefore, the ideal pH range is between 6.0 and 7.8 (^{Safley, 1992}).

We observed a change of EC after 35 d (11.09 to 4.06 mS cm^-1). This result is similar to that reported by ^{Soria et al. (2001)} in bio-digestion of pig liquid manure over a period of 50 d; the process began with 5.8 mS m^-1 and ended with 4.08 mS cm^-1. The authors point out that the decrease in CE was due to the consumption of substrate soluble compounds by microorganisms.This may be possible since microorganisms are in their logarithmic stage of reproduction.

Sodium concentrations in samples from day 1 to 19 were almost constant and an increase was observed from 22 to 48 d, as well as a lower concentration of potassium. This apparently was due to changes in the feed to the digesters.

It is probable that with the decrease and increase of these ions, the average electrical conductivitycwas 8.94± 2.8 and 8.4± 2.71 mS cm^-1 for cattle and porcine bioles. These values are equivalent to their respective sodium, potassium, calcium and magnesium exchangeable bases, whose concentrations also showed higher values in cattle biol and salinity expressed as a percentage of NaCl. Furthermore, the electrical conductivity of very saline soils is between 8 and 16 mS cm^-1; therefore, the biol can be used in low salinity soils or crops resistant to this feature.

Dissolved oxygen (DO) and oxidation reduction potential (ORP)

The DO in cattle and porcine bioles was 0.30± 0.0196 and 0.29± 0.0348, and ORP was -308,8±49.036 and -223,41 ±45,345 mV for each biol (Figure 1). The reduction of dissolved oxygen in relation to water saturated with oxygen (7.21 ppm) showed oxygen consumption by the biodegradation process, and the negative values of ORP indicated that the potential was strongly reducing. The OD and ORP were in line with the anaerobic process in the Sistemas Biobolsa®.

Figure 1 Temperature, pH, electrical conductivity (EC), total dissolved solids (TDS), salinity and dissolved oxygen (DO) in: A) cattle biol; B) porcine biol. 

Humidity percentage, total solids (TS), volatile solids (SSV) and ashes

The average humidity percentages were 95.18 ± 1.90 and 96.21 ± 2.88 %, total solids 4.81 ± 1.89 % and 3.87± 2.89 %, volatile solids 3.29± 1.40 % and 2.48± 2.01 %, and ash 1.51 ± 0.50 % for cattle and porcine bioles, respectively.

The ST in both bioles were lower than those reported by ^{Baserja (1984)} who pointed out that the anaerobic processes with excreta can be unstable, with ST levels below 7 %. Thus, it is possible that the process in our study was within the control limits, 3± s, during the 45-day sampling in both bioles (Figures 1 and 2).

Figure 2 Elemental analysis of micronutrients: A) cattle biol; B) porcine biol discharged by the Sistemas Biobolsa® for two months. 

However, it is possible that the low concentration of ST has affected the poor performance of biogas. In this regard, ^{Budiyono et al. (2010)} evaluated the effect of ST in the speed of biogas production during the anaerobic fermentation of cattle manure, and the optimum range of ST was between 7 and 9 % (which corresponds to a 1:1 excreta and the total volume of liquid ratio). ^{Weiland (2006)} pointed out that in a humid anaerobic fermentation this level is between 8 and 10 %. However, with a low concentration of ST water content increases, and this is one of the most important parameters in the anaerobic digestion due to the movement and growth of bacteria, as well as the easy dissolution and transport of nutrients (^{Sadaka and Engler, 2003}).

Analysis of macro- and micronutrients

The average values of phosphorus and potassium were lower than those reported by ^{Quipuzco et al. (2011)}, and ammonia nitrogen recorded a similar concentration to that observed ^{Pötsch (2004)}. The production of NH₄ - N is due to protein degradation during anaerobic process, and its concentration was higher than any other element. This was consistent with that published by ^{Alvarez and Lide'n (2008)}.

The concentrations of sodium, calcium and magnesium did not change, except on days 18 and 26 (Figure 2). However, the operation of the digester was stable (±3σ variability). The concentrations of these elements were higher than those obtained by ^{Soria et al. (2001)}, which could benefit agricultural crops, when used as a supplement or bio-fertilizer.

The contents of Ca and Mg were significantly (p≤0.05) different between bioles. These differences may be due to diet and digestion (^{Miller and Vare, 2003}), rather than the process itself. Change also depends on the proportion of water and excrement in the food of bio-bags, which the solubilities of the macro and micro elements depend on, which in turn are directly related to the pH (^{López, 1994}; ^{Pérez de Mora et al., 2006}).

Potassium concentration was almost twice that of sodium; this ratio coincided with that obtained by ^{Lamptey et al. (1986)}. Potassium can come in greater proportion from urine rather than manure (^{Yasukawa and Quintero, 1998}).

Organic matter fractions were 32.85 and 23.86 g of L^-1 in cattle and porcine bioles; therefore, they can greatly contribute to improve the quality of soils poor in organic matter.