USA), using as eluant either 0.1 M

USA), using as eluant either 0.1 M sodium phosphate buffer, pH 6.9
(for samples deriving from -amylase hydrolysis) or 0.05 M sodium acetate, pH 6 (for samples deriving from pullulanase hydrolysis), at a flow rate of 0.4 ml/min. The Astra software (ASTRA V 5.1.9.1, Wyatt Technology Co., Santa Barbara, CA, USA) was used for data analysis.


2.5. Pasting properties of starch

Pasting properties were measured in a Brabender Micro-Visco- AmyloGraph (MVAG) (Brabender OHG, Duisburg, Germany). Each sample was finely ground, and 15 g of the resulting powder were dispersed in 100 ml of distilled water. The pasting properties were evaluated, in triplicate, under constant conditions (speed: 250 rpm; sensitivity: 300 cm gf ), according to Marti et al. (2010).

2.6. Properties of the protein network

2.6.1. Protein solubility
Protein solubility in native and denaturing conditions was deter- mined by suspending 0.5 g of finely ground sample in 10 ml of
50 mM phosphate, 0.1 M NaCl, pH 7.0 containing 8 M urea or
8 M urea and 10 mM dithiothreitol (DTT) when indicated. Suspen- sions were stirred for 30 and 60 min at 25 ◦ C. After centrifugation (10,000 × g for 20 min, 20 ◦ C) the amount of protein in the super- natant was determined by a dye-binding method (Bradford, 1976) using bovine serum albumin as a standard. Results are expressed as mg proteins/g sample.


2.6.2. SDS-PAGE
The protein profile in various samples and extraction conditions mentioned above was analysed by SDS-PAGE after denaturation in the presence of 2-mercaptoethanol on a 12% gel using a Miniprotein apparatus (Biorad, Richmond, VA) as described by Cabrera-Chávez et al. (2012). Low molecular weight markers (Amersham Bio- sciences, Amersham, UK) were used for calibration.


2.6.3. Protein thiols
Accessible SH groups were measured by suspending 0.5 g of finely ground sample in 10 ml of 50 mM sodium phosphate buffer, pH 6.8, containing 0.1 M NaCl and 0.2 mM 5,5 -dithiobis(2- nitrobenzoate) (DTNB; Ellman, 1959). After 15 min at room temperature, insoluble material was removed by centrifugation at
12,000 × g for 10 min at 15 ◦ C, and the absorbance at 412 nm of the
supernatant was read against a DTNB blank. Total accessible thiols were measured according to the same protocol outlined above, but adding 8 M urea to the DTNB-containing buffer.



2.7. Cooked pasta characterization

Pasta was cooked in natural spring water (pasta/water ratio
1:10, no salt added) until the optimal cooking time, evaluated according to D’Egidio, Mariani, Nardi, Novaro, & Cubadda, 1990. Optimal cooking times were 9, 10, and 9 min for P1, P2, and P3, respectively. The amount of material leached into cooking water (cooking loss) was measured according to Marti et al. (2010) and expressed as grams of matter lost/100 g of dry pasta. Weight increase of pasta due to water absorption during cooking was eval- uated gravimetrically. Textural characteristics of cooked pasta were determined by using a TA.HD-plus Texture Analyzer (Stable Micro System Ltd., Godalming, UK), equipped with a Kramer cell, accord- ing to Marti et al. (2010). The following indices were considered: compression energy, as the area under the part of the curve related to the compression phase; firmness, as the maximum strength nec- essary to pack the sample; shear force, as the force necessary for

Hasil (Bahasa Indonesia) 1:

USA), menggunakan sebagai eluant baik 0.1 M natrium fosfat buffer, pH 6.9
(untuk sampel berasal dari - amilase hidrolisis) atau 0.05 M natrium asetat, pH 6 (untuk sampel berasal dari hidrolisis pullulanase), dengan laju flow 0.4 ml/menit. Perangkat lunak Astra (ASTRA V 5.1.9.1, Wyatt Teknologi Co, Santa Barbara, CA, USA) digunakan untuk data analisis.


2.5. Menyisipkan sifat Pati

Paste properti diukur dalam Brabender mikro-Visco-AmyloGraph (MVAG) (Brabender OHG, Duisburg, Jerman). Setiap sampel adalah finely tanah, dan 15 g bubuk dihasilkan yang tersebar dalam 100 ml air suling. Sifat paste dievaluasi, dalam rangkap tiga, kondisi konstan (kecepatan: 250 rpm; sensitivitas: gf 300 cm), menurut Marti et al. (2010).

2.6. Sifat-sifat jaringan protein

2.6.1. Kelarutan protein
Protein kelarutan dalam kondisi asli dan mendenaturasikan adalah mencegah - ditambang oleh menangguhkan 0,5 gram sampel tanah finely dalam 10 ml
50 mM fosfat, 0.1 M NaCl pH 7.0 mengandung 8 M urea atau
8 M urea dan 10 mM dithiothreitol (DTT) ketika ditunjukkan. Kandungan Suspen yang diaduk selama 30 dan 60 menit di 25 cita C. Setelah sentrifugasi (10,g × 000 untuk 20 menit, 20 cita C) jumlah protein di super-natant ditentukan oleh mengikat pewarna metode (Bradford, 1976) menggunakan sapi albumin serum sebagai standar. Hasil dinyatakan sebagai mg protein g sampel.


2.6.2. SDS-HALAMAN
Profile protein dalam berbagai sampel dan ekstraksi kondisi diatas dianalisis oleh SDS-halaman setelah denaturasi hadapan 2-mercaptoethanol pada 12% gel menggunakan alat Miniprotein (Biorad, Richmond, VA) seperti yang dijelaskan oleh Cabrera-Chávez et al. (2012). Tanda-tanda berat molekul rendah (Amersham Bio-ilmu, Amersham, Inggris) yang digunakan untuk kalibrasi.


2.6.3. Protein thiols
Diakses SH kelompok diukur oleh menangguhkan 0,5 gram sampel tanah finely dalam 10 ml 50 mM natrium fosfat buffer, pH 6.8, yang mengandung 0.1 M NaCl dan 0.2 mM 5,5 - dithiobis (2-nitrobenzoate) (DTNB; Ellman, 1959). Setelah 15 menit pada suhu kamar, bahan larut dihapus oleh sentrifugasi di
12, 000 g × 10 min di cita 15 C, dan absorbansi di 412 nm
supernatant dibacakan terhadap DTNB kosong. Total thiols diakses diukur sesuai dengan protokol yang sama dijelaskan di atas, tetapi menambahkan 8 M urea yang mengandung DTNB buffer.



2.7. Memasak pasta karakterisasi

Pasta dimasak di mata air alami (pasta air rasio
1:10, tidak ada garam ditambahkan) sampai waktu memasak yang optimal, dievaluasi sesuai dengan D'Egidio, Mariani, Nardi, Novaro, & Cubadda, 1990. Waktu memasak yang optimal adalah 9, 10, dan 9 min untuk P1, P2 dan P3, masing-masing. Jumlah bahan larut ke dalam air memasak (goreng kerugian) diukur menurut Marti et al. (2010) dan dinyatakan sebagai gram masalah kehilangan 100 g pasta kering. Peningkatan berat badan pasta karena penyerapan air selama memasak ini eval-uated gravimetrically. Tekstur karakteristik pasta dimasak ditentukan dengan menggunakan TA.HD-plus tekstur Analyzer (stabil mikro sistem Ltd, Godalming, Inggris), dilengkapi dengan sel Kramer, sesuai-ing untuk Marti et al. (2010). Indeks berikut yang dianggap: kompresi energi, sebagai daerah di bawah bagian dari kurva yang berkaitan dengan fase kompresi; firmness, sebagai kekuatan maksimum nec-essary untuk paket sampel; geser kekuatan, sebagai kekuatan yang diperlukan untuk