Combination Cockle Shells (Anadara granosa) and Calcite Lime to Improve Swamp Water pH for Catfish (Pangasius sp.) Culture Dade

Cockle shells was proven potential used as an alternative liming material. Combination of lime derived from cockle shells with calcite as common lime materials used to overcome limited quantity of cockle shells as the problem of seasonal availability of cockle shells. Besides, these materials have difference characteristics that to be expected can give better effect to pH of soil and water. This study aims to determine the best combination dose between cockle shell limes and calcite to increase the pH of water and soil, survival rate and growth of catfish (Pangasius sp.). The study used completely randomized design (CRD) with 5 treatments and 3 replications. The treatment of different combination of lime doses 100% calcite (P1), 100% cockle shell limes (P2), 75% calcite and 25% cockle shell limes (P3), 50% calcite and 50% cockle shell limes (P4), and 25% calcite and 75% cockle shell limes (P5), dose of lime used as much as 7,000 kg/ha CaO equivalent. The result showed that P4 was the best treatment according to data of swamp water pH was 8.13 and soil pH was 8.07 at final day (day 60), alkalinity 153.33 mg.L CaCO3 equivalent, Ca 104.15 mg.L,100% survival rate, 11.23 cm absolute growth of length, 38.60 g absolute growth of weight and 128.38% feed efficiency.


Introduction
Liming of soil is an effort to increase soil and waters pH of swamp fish ponds. Ponds can be limed with liquid lime, basic slag, or agricultural limestone (Wynne 1996;Boyd 1982Boyd , 1990Wilkinson 2002). Common liming materials include agricultural limestone and liquid lime, calcium hydroxide, calcium oxide and basic slag. Peters (1996) described liming materials that was commonly used are calcitic aglime, dolomitic aglime, hydrated or slaked lime, quicklime or burnt lime, marl, and industrial by product.
An alternative liming material from domestic waste or by-product have an advantage, not only increase pH level, but also environmentally acceptable. The research showed that cockle shells (Anadara granosa) can be used to increase soil and water's pH of swamp fish ponds (Jubaedah et al. 2017). Initial soil's pH value from 3.9 increase to 7.2 as affected by 5 ton/ha lime derived from cockle shells application during 7 days incubation time and maximal water's pH value 7.9 at day 87. Next lime application should be considered after day 205.
According Wilkinson (2002), acid sulfate soils within a pond or its watershed must therefore be treated if the pond is to be used for production. However, large amounts of lime are typically required and the technique may not be economically feasible. Seasonal availability of cockle shells needs to combine with common agriculture limestone.
Calcite (Calcium carbonates, CaCO3) is one kind of agricultural lime stone that slowly react but simultaneously safe to fish (Boyd, 1982;1990) Calcium carbonates on cockle shells converted to calcium oxide through calcination process. CaO short-term acting but can be unsafe for fish (Nobre et al. 2014). The present work aimed to compare the pH of soil and water as well as survival rate and growth performance of catfish juvenile reared in ponds with application of combination of combination of calcite (CaCO3) and lime derived from cockle shells.

Preparing Lime Materials and Liming Application
Calcination process of A. granosa shells according to Jubaedah et al. (2017), including burning process using furnace at temperature of 800 o C for an hour. The calcin then mashed and sieved with 60, 40 and 20 mesh-size sieves in order to get lime with 50 % passed 60 mesh-size sieve, 25 % passed 40 mesh-sized sieve, and 25 % passed 20 mesh-size sieve.
Five combination of lime were investigated, namely: P1) 100% calcite, P2) 100% cockle shells lime, P3) 75% calcite and 25% cockle shell limes, P4) 50% calcite and 50% cockle shell limes, and P5) 25% calcite and 75% cockle shell limes, dose of lime each used as much 7,000 kg/ha CaO equivalent. Lime were applied homogeneously on soil ponds and incubated for 7 days at field capacity soil moisture. Water was filled to the ponds and and let equilibrated for 3 days. Fish stocked on ponds and cultured for 60 days.

Fish Culture
Fifteen unit of fish ponds filled swamp water with 500 L/ponds at 7 days after liming. Fish were acclimatized for one week before used in this research. The fifteens of 5 + 0.5 cm length Pangasius sp. stocked for each pond.
Starting from 10 days after liming, fish was cultured for 60 days. The fish were fed to satiation three times per day with an artificial diet containing 30% protein. Water samples were collected and analyzed, as well as growth of fish was measured every 20 days.

Experimental variables and analytical procedures
Water quality, survival and growth performance variables were observed in the present work. The water's pH, total alkalinity, and total ammonia were monitored every 20day, Ca concentration analyzed every 30 days in all pond. Besides, water temperature and pH were recorded daily. The water pH was measured by using a portable pH meter. The water temperature was observed by using a digital handy thermometer. The analytical determinations of total alkalinity and total ammonia and Ca were carried out according to the guidelines presented by APHA (2012). The fish initial and final body weight and length, survival and feed efficiency were observed in all experimental units.
Water quality, survival and growth performance results were statistically analyzed according to the two-way Anova to detect if there was any significant influence due to experimental treatments. When the influence was at least significant, the means were compared using Least Significant Difference (LSD) test.

Data Analysis
The fish survival rate was calculated from the initial number of fish and mortality after the experiment was completed. The absolute growth of fish was determined from the mean of initial and final weight and length of fish, respectively for absolute weight growth and length growth. Meanwhile, feed efficiency was calculated by the formula of NRC (1977).

Results and Discussion
The initial Soil pH used in this current study was 3.60 and then soil pH increased due to liming to pH 7.40-7.53 after incubated for 7 days (Figure 1). The soil's pH tends to increase rapidly until neutral pH value at the beginning of incubation, then increase slowly.
Regression between combination of limes and pH of soil and water showed polynomial quadratic patterns for day 0, 10, 20 and 30 (Figure 2 and Figure 3). The results showed combination of calcite (50%) and cockle shells lime (50%) has highest soil's and water's pH. The calcination process of cockle shells converts calcium carbonate (CaCO3) to calcium oxide (CaO). According to Boyd (1982;1990), reaction with acidity is faster than CaCO3, but highly caustic and cause the water pH raise to levels toxic for fish. Calcite (CaCO3) is agricultural limestone that is slow-acting product but generally the safest (Boyd 1982;Nobre et al. 2014). The combination of cockle shells lime and calcite raise pH value higher than single lime, both cockle shells and calcite.
The initial water pH used in this research was 3.5, then water pH increased following polynomial quadratic patterns for 60 days of fish culture due to lime application (Figure 4). According to equations shown in Figure 3, the maximum pH and days after lime application to reach maximum water pH can be calculated. The maximum water pH and when it is were 7.85 at 50 days after liming, 7.82 at 50 days after liming, 7.93 at 50 days after liming, 8.31 at 89 days after liming and 8.03 at 59 days after liming, respectively for P1, P2, P3, P4 and P5.
The highest maximum water pH among treatments was observed on P4 (50% calcite and 50% cockle shell lime). Furthermore, based on the equations (Figure 4) and minimum water pH (6.5) for optimal growth of Pangasius sp., the next lime application should be considered after day, 132 th , 132 th , 134 th , 224 th and 146 th for P1, P2, P3, P4 and P5, respectively.
Liming not only increase pH value, but also alkalinity of water. Based on Figure 5 showed that liming application of combination cockle shell limes and calcite affected to increase alkalinity of water. Greater alkalinity after liming also buffer water from drastic daily changes in pH (Boyd, 1990).
Liming also related to calcium (Ca) concentration. Fish will lose Ca to the water on extremely low Ca concentration in the water, even though the minimum acceptable Ca concentration in the water necessary to permit optimum fish growth has not been established (Boyd, 1990). The liming was giving significantly different to Ca concentration at final day of research (day 60 th ) ( Table 1). Combination of cockle shell lime 50% and calcite 50% (P4) has   highest Ca, significance different with calcite 100% (P1) and cockle shell 100% (P2), but no significant different with combination cockle shell lime 75% calcite and 25% cockle shell limes (P3), and 25% calcite and 75% cockle shell limes (P5). According to Boyd et al. (2002), Ca concentration from calcite is 40.08%, meanwhile based on the analysis of Ca content from cockle shells is 61.16%. The water temperature and dissolved oxygen (DO) of ponds (Table 2) ranged from 25.3 to 32.2 o C and from 5.4 to 6.6 mg.L -1 , respectively. Based on National standardization Agency or Badan Standar Nasional Indonesia (BSNI, 2000), the temperature of water was exceed optimum ranged that is 27-30 o C according to optimal value of temperature for Pangasius sp. but still in tolerance ranged for fish. Meanwhile, the DO was approriate for Pangasius sp. No significant effect of lime application on dissolved oxygen at day 0, 20,40, and 60 of the rearing period.
All treatment showed that the ammonia concentration increased in the final day of research (day 60 th of culture) (Figure 6). The ammonia concentration at culture periods under maximal value for amonia for fish according to Indonesian environmental regulation or Peraturan Pemerintah Republik Indonesia (2001) that is >1 mg.L -1 .
The survival rate of Pangsius sp. Were 100% for all treatments. The liming application using single lime materials both calcite 100% and cockle shells 100%, or combination of them can increase pH until optimum for fish. In general, the water quality supported for living of fish.
The growth performance and fed efficiency showed at Table 3 Based on analysis of variance showed that treatment gave an effect to growth performance of fish. The highest absolute growth (length and weight) of fishes was on combination 50% cockle shells and 50% calcite (P4) and significant difference with others.
The highest feed efficiency was on fish cultured at ponds with liming combination calcite (50%) and cockle shells (50%). It is probably related to availability of plankton as natural feed of fish. The highest alkalinity shown at P4 ( Figure  5) and it will increase the carbon dioxide for photosynthesis of phytoplankton. According to Boyd (1982Boyd ( , 1990 and Wilkinson (2002), the positive effect of liming was increasing the alkalinity of water, thereby increasing the    availability of carbon dioxide for phytoplankton growth. Wilkinson (2002), from some references conclude that applying lime has been shown to increase soluble phosphorus concentrations as nutrient of phytoplankton. Lime provides calcium and neutralizes acidic soils so that phosphate is released as calcium exchanges with aluminum and iron. Calcium phosphates are more soluble in water than aluminum phosphates or iron phosphates (Burtle, 2015).

Conclusion
The combination of 50% cockle shells lime and 50% calcite was the best treatments with pH of soil and water at day 60 were 8.07 and 8.31, respectively; highest maximal water pH value (3.31) and next liming was at day 224; highest alkalinity (153.33 mg.L -1 CaCO3 equivalent), and Ca (104.15 mg.L -1 ); highest feed efficiency (128.38%); and the best growth performance. The water quality including water temperature, dissolved oxygen and ammonia were in appropriate for Pangasius sp. with 100% survival rate.